Characterization of Oogonial Stem Cells in Adult Mouse Ovaries with Age and Comparison to In Silico Data on Human Ovarian Aging

Abstract

Many adult somatic stem cell lineages are comprised of subpopulations that differ in gene expression, mitotic activity, and differentiation status. In this study, we explored if cellular heterogeneity also exists within oogonial stem cells (OSCs), and how chronological aging impacts OSCs. In OSCs isolated from mouse ovaries by flow cytometry and established in culture, we identified subpopulations of OSCs that could be separated based on differential expression of stage-specific embryonic antigen 1 (SSEA1) and cluster of differentiation 61 (CD61). Levels of aldehyde dehydrogenase (ALDH) activity were inversely related to OSC differentiation, whereas commitment of OSCs to differentiation through transcriptional activation of stimulated by retinoic acid gene 8 was marked by a decline in ALDH activity and in SSEA1 expression. Analysis of OSCs freshly isolated from ovaries of mice between 3 and 20 months of age revealed that these subpopulations were present and persisted throughout adult life. However, expression of developmental pluripotency associated 3 ( Dppa3 ), an epigenetic modifier that promotes OSC differentiation into oocytes, was lost as the mice transitioned from a time of reproductive compromise (10 months) to reproductive failure (15 months). Further analysis showed that OSCs from aged females could be established in culture, and that once established the cultured cells reactivated Dppa3 expression and the capacity for oogenesis. Analysis of single-nucleus RNA sequence data sets generated from ovaries of women in their 20s versus those in their late 40s to early 50s showed that the frequency of DPPA3 -expressing cells decreased with advancing age, and this was paralleled by reduced expression of several key meiotic differentiation genes. These data support the existence of OSC subpopulations that differ in gene expression profiles and differentiation status. In addition, an age-related decrease in Dppa3 / DPPA3 expression, which is conserved between mice and humans, may play a role in loss of the ability of OSCs to maintain oogenesis with age.

Pluripotent stem cells: Basic biology or else differentiations aimed at translational research and the role of flow cytometry

“…Such hepatocytes have been proven to be more robust than those differentiated from iPSC [25]. In a different context than the PSC, other stem cells, such as oogonial stem cells (OSC), have been analyzed at a single cell level using flow cytometry for precise characterization [26]. OSCs harbor immense heterogeneity and the cell types in OSCs help one understand their respective differentiation potential into oocytes [26].…”

“…In a different context than the PSC, other stem cells, such as oogonial stem cells (OSC), have been analyzed at a single cell level using flow cytometry for precise characterization [26]. OSCs harbor immense heterogeneity and the cell types in OSCs help one understand their respective differentiation potential into oocytes [26]. For example, subpopulations of OSCs expressed SSEA‐1, cluster of differentiation 61(CD61), aldehyde dehydrogenase (ALDH), and developmental pluripotency associated 3 (Dppa3).…”

“…ALDH and SSEA1 subpopulations existed in OSCs in all ages of mice. However, the Dppa3 (an epigenetic modifier) showed a decline from 10 months (reproductive compromise) to 15 months (reproductive failure) in mice, thereby indicating the importance of single‐cell analysis for downstream applications [26]. Such data in mice was corroborated in human females showing a reduced Dppa3 activity in ovaries at ages the 40s to 50s, indicating a gradual reproductive failure [26].…”

“…Hence, it is advisable to sort the small populations of mesenchymal stem cells residing in the SVF and reprogram them into iPSC or lineage reprogramming into a specific lineage, for example, hepatocytes [25]. Such hepatocytes have been proven to be more robust than those differentiated from iPSC [25].In a different context than the PSC, other stem cells, such as oogonial stem cells (OSC), have been analyzed at a single cell level using flow cytometry for precise characterization [26].…”

“…OSCs harbor immense heterogeneity and the cell types in OSCs help one understand their respective differentiation potential into oocytes [26]. For example, subpopulations of OSCs expressed SSEA-1, cluster of differentiation 61(CD61), aldehyde dehydrogenase (ALDH), and developmental pluripotency associated 3 (Dppa3).…”