A now large body of evidence supports the existence of mitotically active germ cells in postnatal ovaries of diverse mammalian species, including humans. This opens the possibility that adult stem cells naturally committed to a germline fate could be leveraged for the production of female gametes outside of the body. The functional properties of these cells, referred to as female germline or oogonial stem cells (OSCs), in ovaries of women have recently been tested in various ways, including a very recent investigation of the differentiation capacity of human OSCs at a single cell level. The exciting insights gained from these experiments, coupled with other data derived from intraovarian transplantation and genetic tracing analyses in animal models that have established the capacity of OSCs to generate healthy eggs, embryos and offspring, should drive constructive discussions in this relatively new field to further exploring the value of these cells to the study, and potential management, of human female fertility. Here, we provide a brief history of the discovery and characterization of OSCs in mammals, as well as of the in-vivo significance of postnatal oogenesis to adult ovarian function. We then highlight several key observations made recently on the biology of OSCs, and integrate this information into a broader discussion of the potential value and limitations of these adult stem cells to achieving a greater understanding of human female gametogenesis in vivo and in vitro.

Mitochondria are well-characterized regarding their function in both energy production and regulation of cell death; however, the heterogeneity that exists within mitochondrial populations is poorly understood. Typically analyzed as pooled samples comprised of millions of individual mitochondria, there is little information regarding potentially different functionality across subpopulations of mitochondria. Herein we present a new methodology to analyze mitochondria as individual components of a complex and heterogeneous network, using a nanoscale and multi–parametric flow cytometry-based platform. We validate the platform using multiple downstream assays, including electron microscopy, ATP generation, quantitative mass-spectrometry proteomic profiling, and mtDNA analysis at the level of single organelles. These strategies allow robust analysis and isolation of mitochondrial subpopulations to more broadly elucidate the underlying complexities of mitochondria as these organelles function collectively within a cell.

Contrasting the equal contribution of nuclear genetic material from maternal and paternal sources to offspring, passage of mitochondria, and thus mitochondrial DNA (mtDNA), is uniparental through the egg. Since mitochondria in eggs are ancestral to all somatic mitochondria of the next generation and to all cells of future generations, oocytes must prepare for the high energetic demands of maturation, fertilization and embryogenesis while simultaneously ensuring that their mitochondrial genomes are inherited in an undamaged state. Although significant effort has been made to understand how the mtDNA bottleneck and purifying selection act coordinately to prevent silent and unchecked spreading of invisible mtDNA mutations through the female germ line across successive generations, it is unknown if and how somatic cells of the immediate next generation are spared from inheritance of detrimental mtDNA molecules. Here, we review unique aspects of mitochondrial activity and segregation in eggs and early embryos, and how these events play into embryonic developmental competency in the face of advancing maternal age.

Multiple labs have reported that mammalian ovaries contain oogonial stem cells (OSCs), which can differentiate into oocytes that fertilize to produce offspring. However, the physiological relevance of these observations to adult ovarian function is unknown. Here we performed targeted and reversible ablation of premeiotic germ cells undergoing differentiation into oocytes in transgenic mice expressing the suicide gene, herpes simplex virus thymidine kinase (HSVtk), driven by the promoter of stimulated by retinoic acid gene 8 (Stra8), a germ cell-specific gene activated during meiotic commitment. Over a 21-day ablation phase induced by the HSVtk pro-drug, ganciclovir (GCV), oocyte numbers declined due to a disruption of new oocyte input. However, germ cell differentiation resumed after ceasing the ablation protocol, enabling complete regeneration of the oocyte pool. We next employed inducible lineage tracing to fate map, through Cre recombinase-mediated fluorescent reporter gene activation only in Stra8-expressing cells, newly-formed oocytes. Induction of the system during adulthood yielded a mosaic pool of unmarked (pre-existing) and marked (newly-formed) oocytes. Marked oocytes matured and fertilized to produce offspring, which grew normally to adulthood and transmitted the reporter to second-generation offspring. These findings establish that oocytes generated during adulthood contribute directly to ovarian function and natural fertility in mammals.

Historically, approaches designed to offer women diagnosed with cancer the prospects of having a genetically matched child after completion of their cytotoxic treatments focused on the existing oocyte population as the sole resource available for clinical management of infertility. In this regard, elective oocyte and embryo cryopreservation, as well as autologous ovarian cortical tissue grafting posttreatment, have gained widespread support as options for young girls and reproductive-age women who are faced with cancer to consider. In addition, the use of ovarian protective therapies, including gonadotropin-releasing hormone agonists and sphingosine-1-phosphate analogs, has been put forth as an alternative way to preserve fertility by shielding existing oocytes in the ovaries in vivo from the side-effect damage caused by radiotherapy and many chemotherapeutic regimens. This viewpoint changed with the publication of now numerous reports that adult ovaries of many mammalian species, including humans, contain a rare population of oocyte-producing germ cells—referred to as female germline or oogonial stem cells (OSCs). This new line of study has fueled research into the prospects of generating new oocytes, rather than working with existing oocytes, as a novel approach to sustain or restore fertility in female cancer survivors. Here, we overview the history of work from laboratories around the world focused on improving our understanding of the biology of OSCs and how these cells may be used to reconstitute “artificial” ovarian tissue in vitro or to regenerate damaged ovarian tissue in vivo as future fertility-preservation options.