Publications

Noninvasive index of cryorecovery and growth potential for human follicles in vitro.

Noninvasive index of cryorecovery and growth potential for human follicles in vitro.

Biol Reprod. 2010 Jun;82(6):1180-9

Authors: Barrett SL, Shea LD, Woodruff TK

Abstract

Cryopreservation of oocytes and embryos is commonly used to preserve fertility. However, women undergoing cancer treatment may not have the time or may not be good candidates for these options. Ovarian cortical tissue cryopreservation and subsequent tissue transplant has been proven successful yet inefficient in preserving larger secondary follicles, and is not recommended as a fertility preservation option for women with certain cancers. We evaluated cryopreservation of individual follicles as an alternative option in rodents, nonhuman primates, and human primates. Under optimal conditions, cryopreserved mouse secondary follicles were able to reestablish granulosa cell-oocyte interactions, which are essential for subsequent follicle growth. Individual secondary follicles survived cryopreservation, were able to be cultured in a three-dimensional alginate hydrogel matrix to the antral stage, and the enclosed oocytes were competent for fertilization. Using a vital imaging technique (pol-scope) employed in many fertility centers, we were able to bioassay the thawed, cultured follicles for the presence of transzonal connections between the somatic and germ cells. Perturbations in these linkages were shown to be reversed when follicles were cryopreserved under optimal freezing conditions. We applied the optimized cryopreservation protocol to isolated rhesus monkey and human secondary follicles, and using the birefringent bioassay, we were able to show good correlation between early follicle growth and healthy somatic cell-oocyte connections. Our results suggest that ovarian follicles can be cryopreserved, thawed, and analyzed noninvasively, making follicle preservation an additional option for young cancer patients.

PMID: 20200211 [PubMed - indexed for MEDLINE]

The time is now for a new approach to primary ovarian insufficiency.

The time is now for a new approach to primary ovarian insufficiency.

Fertil Steril. 2011 May;95(6):1890-7

Authors: Cooper AR, Baker VL, Sterling EW, Ryan ME, Woodruff TK, Nelson LM

Abstract

OBJECTIVE: To articulate the need for a new approach to primary ovarian insufficiency. The condition, also known as premature menopause or premature ovarian failure, is defined by the presence of menopausal-level serum gonadotropins in association with irregular menses in adolescent girls or women younger than 40 years. It can be iatrogenic as related to cancer therapy or may arise spontaneously, either alone or as part of a host of ultrarare syndromes. In a large percentage of spontaneous cases no pathogenic mechanism can be identified.

DESIGN: Literature review and consensus building at a multidisciplinary scientific workshop.

CONCLUSION(S): There are major gaps in knowledge regarding the etiologic mechanisms, psychosocial effects, natural history, and medical and psychosocial management of primary ovarian insufficiency. An international research consortium and disease registry formed under the guidance of an umbrella organization would provide a pathway to comprehensively increase basic and clinical knowledge about the condition. Such a consortium and patient registry also would provide clinical samples and clinical data with a goal toward defining the specific pathogenic mechanisms. An international collaborative approach that combines the structure of a patient registry with the principles of integrative care and community-based participatory research is needed to advance the field of primary ovarian insufficiency.

PMID: 20188353 [PubMed - indexed for MEDLINE]

The Illinois Women's Health Registry: advancing women's health research and education in Illinois, USA.

The Illinois Women's Health Registry: advancing women's health research and education in Illinois, USA.

Womens Health (Lond Engl). 2010 Mar;6(2):183-96

Authors: Bristol-Gould S, Desjardins M, Woodruff TK

Abstract

To achieve the goal of personalized medicine, we must first improve our understanding of the differences in health and illness between men and women. The purpose of the Illinois Women's Health Registry (USA) is to provide a research and education tool that advances scientific knowledge of sex- and gender-based differences in health and disease. Specifically, the Registry is a confidential 30-min health and lifestyle survey for female residents of Illinois over the age of 18 years. The survey includes questions regarding health, environment, health-related behaviors, symptoms and illnesses or conditions that a participant may have now or has had in the past. By enrolling in the Registry, women throughout the state are provided with information and access to clinical research studies that they may be eligible for, based on their self-reported health information. The Registry not only serves as a platform for recruitment into pivotal research studies, but also represents the beginning of a state-wide database that enables researchers to examine the collective de-identified health information provided by women living in Illinois. Ultimately, a cross-sectional and longitudinal analysis of these data will help to clarify the issues that women themselves identify as their main health concerns. In response to these concerns, specific research studies can be designed and launched, allowing us to eventually deliver tailored treatment and prevention options to women. Finally, by creating a reliable state-focused research tool, developed by staff that are trained in women's health research, we can compare health issues across the state and apply strategies for improvement where it is needed most. This article will provide examples of sex differences in disease, the lack of federal enforcement for inclusion of women in studies, researcher-perceived burdens and sex-based reasons as to why recruitment of women is considered to be more challenging. In addition, this article will discuss what a women's health registry is and why we need one in Illinois, how we have recruited women and our successes and challenges. Our goal is to inform the reader about the utility of a state-based tool and to provide a discussion regarding the lessons learned in order to aid other states in implementing this kind of program.

PMID: 20187725 [PubMed - indexed for MEDLINE]

Altered sex hormone concentrations and gonadal mRNA expression levels of activin signaling factors in hatchling alligators from a contaminated Florida lake.

Altered sex hormone concentrations and gonadal mRNA expression levels of activin signaling factors in hatchling alligators from a contaminated Florida lake.

J Exp Zool A Ecol Genet Physiol. 2010 Apr 1;313(4):218-30

Authors: Moore BC, Kohno S, Cook RW, Alvers AL, Hamlin HJ, Woodruff TK, Guillette LJ

Abstract

Activins and estrogens participate in regulating the breakdown of ovarian germ cell nests and follicle assembly in mammals. In 1994, our group reported elevated frequencies of abnormal, multioocytic ovarian follicles in 6 month old, environmental contaminant-exposed female alligators after gonadotropin challenge. Here, we investigated if maternal contribution of endocrine disrupting contaminants to the egg subsequently alters estrogen/inhibin/activin signaling in hatchling female offspring, putatively predisposing an increased frequency of multioocytic follicle formation. We quantified basal and exogenous gonadotropin-stimulated concentrations of circulating plasma steroid hormones and ovarian activin signaling factor mRNA abundance in hatchling alligators from the same contaminated (Lake Apopka) and reference (Lake Woodruff) Florida lakes, as examined in 1994. Basal circulating plasma estradiol and testosterone concentrations were greater in alligators from the contaminated environment, whereas activin/inhibin betaA subunit and follistatin mRNA abundances were lower than values measured in ovaries from reference lake animals. Challenged, contaminant-exposed animals showed a more robust increase in plasma estradiol concentration following an acute follicle stimulating hormone (FSH) challenge compared with reference site alligators. Aromatase and follistatin mRNA levels increased in response to an extended FSH challenge in the reference site animals, but not in the contaminant-exposed animals. In hatchling alligators, ovarian follicles have not yet formed; therefore, these endocrine differences are likely to affect subsequent ovarian development, including ovarian follicle assembly.

PMID: 20166196 [PubMed - indexed for MEDLINE]

Current achievements and future research directions in ovarian tissue culture, in vitro follicle development and transplantation: implications for fertility preservation.

Current achievements and future research directions in ovarian tissue culture, in vitro follicle development and transplantation: implications for fertility preservation.

Hum Reprod Update. 2010 Jul-Aug;16(4):395-414

Authors: Smitz J, Dolmans MM, Donnez J, Fortune JE, Hovatta O, Jewgenow K, Picton HM, Plancha C, Shea LD, Stouffer RL, Telfer EE, Woodruff TK, Zelinski MB

Abstract

BACKGROUND Female cancer patients are offered 'banking' of gametes before starting fertility-threatening cancer therapy. Transplants of fresh and frozen ovarian tissue between healthy fertile and infertile women have demonstrated the utility of the tissue banked for restoration of endocrine and fertility function. Additional methods, like follicle culture and isolated follicle transplantation, are in development. METHODS Specialist reproductive medicine scientists and clinicians with complementary expertise in ovarian tissue culture and transplantation presented relevant published literature in their field of expertise and also unpublished promising data for discussion. As the major aims were to identify the current gaps prohibiting advancement, to share technical experience and to orient new research, contributors were allowed to provide their opinioned expert views on future research. RESULTS Normal healthy children have been born in cancer survivors after orthotopic transplantation of their cryopreserved ovarian tissue. Longevity of the graft might be optimized by using new vitrification techniques and by promoting rapid revascularization of the graft. For the in vitro culture of follicles, a successive battery of culture methods including the use of defined media, growth factors and three-dimensional extracellular matrix support might overcome growth arrest of the follicles. Molecular methods and immunoassay can evaluate stage of maturation and guide adequate differentiation. Large animals, including non-human primates, are essential working models. CONCLUSIONS Experiments on ovarian tissue from non-human primate models and from consenting fertile and infertile patients benefit from a multidisciplinary approach. The new discipline of oncofertility requires professionalization, multidisciplinarity and mobilization of funding for basic and translational research.

PMID: 20124287 [PubMed - indexed for MEDLINE]

Markers of growth and development in primate primordial follicles are preserved after slow cryopreservation.

Markers of growth and development in primate primordial follicles are preserved after slow cryopreservation.

Fertil Steril. 2010 May 15;93(8):2627-32

Authors: Jin S, Lei L, Shea LD, Zelinski MB, Stouffer RL, Woodruff TK

Abstract

OBJECTIVE: To investigate the effect of slow cryopreservation on the morphology and function of primate primordial follicles within ovarian tissue slices.

DESIGN: Fresh monkey ovarian tissue was frozen by slow cryopreservation and thawed for analysis of morphologic and functional parameters.

SETTING: University-affiliated laboratory.

ANIMALS: Rhesus monkey ovarian tissue.

INTERVENTION(S): None.

MAIN OUTCOME MEASURE(S): Histologic analysis, follicle counting, assessment of protein abundance and localization.

RESULT(S): After freezing and thawing, 89% of the primordial follicles maintained their laminar-based architecture, with sizes close to those of fresh fixed follicles. Molecular markers of early follicle health (activin subunits and the phosphorylated form of the signaling protein Smad2 [pSmad2]) were present in fresh and frozen-thawed primordial follicles. Stroma cells, but not follicles, had a higher level of TUNEL staining. Granulosa cells within the follicles of frozen-thawed ovarian tissue cultured for 48 hours had the capacity to proliferate and sustained expression of the activin subunits and nuclear pSmad2.

CONCLUSION(S): This study provides evidence that markers of early follicle growth and development are preserved after slow cryopreservation and thaw, with little effect on follicle morphology and function.

PMID: 20074723 [PubMed - indexed for MEDLINE]

Rock the cradle.

Rock the cradle.

Nat Med. 2009 Nov;15(11):1244

Authors: Woodruff TK

PMID: 19893551 [PubMed - in process]

Preserving fertility during cancer treatment.

Preserving fertility during cancer treatment.

Nat Med. 2009 Oct;15(10):1124-5

Authors: Woodruff TK

PMID: 19812566 [PubMed - indexed for MEDLINE]

The regulatory role of Dicer in folliculogenesis in mice.

The regulatory role of Dicer in folliculogenesis in mice.

Mol Cell Endocrinol. 2010 Feb 5;315(1-2):63-73

Authors: Lei L, Jin S, Gonzalez G, Behringer RR, Woodruff TK

Abstract

Dicer is the ribonuclease III for synthesis of mature functional microRNAs (miRNAs), which play an important role in regulating cell development. In the mouse ovary, the Dicer1 protein was expressed in both oocyte and granulosa cells of the follicle. In the present study, the role of miRNAs in mouse ovarian development was explored by using Dicer1 conditional knockout (cKO) mouse ovarian tissue (Amhr2 Cre/-; Dicer flox/flox), in which Dicer1 is deleted specifically in follicular granulosa cells. The morphology and gene expression profile of cKO and wild type (WT) mouse ovaries at various stages of development (day 4, day 8, 8 weeks and 8 months) were examined. Comparative analysis of the follicle number indicated that conditional inactivation of Dicer1 in the follicular granulosa cells led to an increased primordial follicle pool endowment, accelerated early follicle recruitment and more degenerate follicles in the cKO ovaries. In addition, significant differences were noted in the expression of some follicle development-related genes between cKO and WT mouse ovaries, such as Amh, Inhba, Cyp17a1, Cyp19a1, Zps, Gdf9 and Bmp15, suggesting the function of miRNAs in regulating gene expression is time- and gene-dependent. With the Dicer1 inactivation, mmu-mir-503, a miRNA that is more abundant in mouse ovary than in other tissues, was down-regulated significantly. Meanwhile, the expression of mmu-mir-503 decreased notably with follicle development in the gonadotropin-primed mouse ovary. Up-regulation of mmu-mir-503 in primary cultured granulosa cells resulted in the decreased expression of both the target gene and non-target gene at the transcriptional level, which involve genes related to granulosa cell proliferation and luteinization. In conclusion, Dicer1 plays important roles in follicular cell development through the differential regulation of gene expression.

PMID: 19799966 [PubMed - indexed for MEDLINE]

The primordial pool of follicles and nest breakdown in mammalian ovaries.

The primordial pool of follicles and nest breakdown in mammalian ovaries.

Mol Hum Reprod. 2009 Dec;15(12):795-803

Authors: Tingen C, Kim A, Woodruff TK

Abstract

The creation of the pool of follicles available for selection and ovulation is a multi-faceted, tightly regulated process that spans the period from embryonic development through to the first reproductive cycle of the organism. In mice, this development can occur in mere weeks, but in humans, it is sustained for years. Embryonic germ cell development involves the migration of primordial germs cells to the genital ridge, and the mitotic division of germ cell nuclei without complete cytokinesis to form a multi-nucleated syncytia, or germ cell nest. Through combined actions of germ cell apoptosis and somatic cell migration, the germ cell nuclei are packaged, with surrounding granulosa cells, into primordial follicles to form the initial follicle pool. Though often dismissed as quiescent and possibly uninteresting, this initial follicle pool is actually quite dynamic. In a very strictly controlled mechanism, a large portion of the initial primordial follicles formed is lost by atresia before cycling even begins. Remaining follicles can undergo alternate fates of continued dormancy or selection leading to follicular growth and differentiation. Together, the processes involved in the fate decisions of atresia, sustained dormancy, or activation carve out the follicle pool of puberty, the pool of available oocytes from which all future reproductive cycles of the female can choose. The formation of the initial and pubertal follicle pools can be predictably affected by exogenous treatment with hormones or molecules such as activin, demonstrating the ways the ovary controls the quality and quantity of germ cells maintained. Here, we review the biological processes involved in the formation of the initial follicle pool and the follicle pool of puberty, address the alternate models for regulating germ cell number and outline how the ovary quality-controls the germ cells produced.

PMID: 19710243 [PubMed - indexed for MEDLINE]