The n e w e ng l a n d j o u r na l of m e dic i n e

Cl inic a l I m pl ic a t ions of B a sic R e se a rch

Elizabeth G. Phimister, Ph.D., Editor

Oocytes from Stem Cells

Mary Herbert, Ph.D., and Azim Surani, Ph.D.

Primordial germ cells, which emerge around the primordial germ cells proliferate before ma-
day 16 in early human embryos, develop to form turing to form oogonia.2 These cells then enter
oogonia so that eventually, the number of oocytes into meiosis and eventually develop as oocytes.
that are present in an adult female is fixed at Meiosis ensures that each female gamete is hap-
birth. After undergoing a complex developmental loid and thus will complement the equivalent pa-
program in fetal gonads, primordial germ cells ternal contribution on fertilization. Meiosis and
develop to form oogonia, which form oocytes oocyte formation are genetically separable events,3
capable of undergoing maturation and fertiliza- but faulty meiosis is a barrier to the development
tion (Fig. 1). To recapitulate the entire female of functional oocytes capable of transmitting a
germline life cycle with stem cells and end with single copy of each chromosome.
functional oocytes in culture is a daunting task Hayashi and colleagues generated ovarian
but nonetheless one that was recently accom- somatic cells from mouse embryonic stem cells
plished by Yoshino, Hayashi, and colleagues.1 in culture by mimicking stepwise the develop-
Starting with mouse embryonic stem cells, these mental trajectories from mesoderm to the fetal
investigators generated fetal ovarian somatic ovarian stroma, granulosa, and theca cells that
cells, which supported the development of pri- constitute the ovarian follicle. The aggregation
mordial germ-cell–like cells (also derived from of embryonic stem-cell–derived ovarian somatic
embryonic stem cells) into oocytes that were cell–like cells with embryonic stem-cell–derived
capable of being fertilized. A subset of embryos primordial germ-cell–like cells resulted in recon-
developed to term. stituted ovarioids. By day 3 of culture, granulosa
Primordial germ cells and gonads develop in- cells surrounded the primordial germ-cell–like
dependently in embryos, so primordial germ cells cells, promoting their proliferation and expres-
are required to migrate to the developing go- sion of the late germ-cell marker DDX4. Zipper-
nads; this migration is followed by synchronized like structures called synaptonemal complexes,
development of the two entities, during which in which paired homologous chromosomes

Figure 1 (facing page). Recapitulation of Ovarian Somatic Cell and Germ-Cell Development in the Generation of Functional Oocytes
from Pluripotent Stem Cells.
Primordial germ cells (PGCs) migrate to the developing gonads and proliferate. Gonadal somatic cells differentiate into ovarian stroma,
granulosa, and theca cells. In parallel, germ cells differentiate and enter meiosis, forming oocytes, which, surrounded by a layer of gran-
ulosa cells, form primordial follicles. After recruitment for growth, primordial follicles develop to the secondary follicle stage, containing
multiple layers of granulosa cells surrounded by theca cells. In sexually mature females, follicles develop into the preovulatory stage and
oocytes resume meiosis, arresting after completion of the first meiotic division before ovulation (Panel A). In the in vitro differentiation
of ovarian somatic cells and PGCs from embryonic stem cells (Panel B), the aggregation of embryonic stem-cell–derived fetal ovarian
somatic–like cells with PGC-like cells promotes the proliferation and differentiation of both lineages to form reconstituted ovarioids. The
follicle-like structures in the ovarioid progress from the primordial follicle stage to the secondary follicle stage. Transferred to conditions
that promote growth, the secondary follicle–like structures develop to form preovulatory-like follicles, from which cumulus–oocyte com-
plexes are harvested and matured in vitro. Fertilization of the mature oocytes results in viable embryos.

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 Clinical Implications of Basic Research

Proliferation of PGCs Meiosis Primordial follicles

FETAL OVARY NEONATAL OVARY

Primordial
Granulosa follicles
Theca
cell cell
Secondary Primary
Cumulus–oocyte follicle follicle
complex
follicle

Migration of PGCs to fetal Preovulatory

gonadal ridge follicle

Ovulating ADULT OVARY

follicle

B Differentiation to fetal
ovarian somatic Aggregation Reconstituted
cell–like cells ovarioid

Mouse
pluripotent
stem cells

Differentiation to
PGC-like cells

Cumulus–oocyte
complex Theca cell
Blastocyst Granulosa
cell

Offspring
2-cell embryo Mature oocyte Oocyte

Fertilization Preovulatory Antral Secondary

In vitro maturation In vitro growth of follicle-like structures

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 Clinical Implications of Basic Research

undergo meiotic recombination,4 were evident in Investigations into the genesis of human
germ-cell nuclei by day 7. As the cultures pro- oogonia and oocytes are challenging because
gressed, follicle-like structures became surround- of the inaccessibility of the gonads, so in vitro
ed by a layer of theca-like cells. After 23 days of models are essential for studying the otherwise
culture, oocytes were surrounded by multiple intractable early stages of germ-cell development
layers of granulosa-like cells to form secondary and oogenesis. For example, what governs the
follicle–like structures, which were then placed positioning of meiotic recombination sites that
in specific culture conditions that have previously subsequently render female meiosis susceptible
been shown to support growth to the preovula- to chromosome missegregation? An experimen-
tory stage.5 During 12 days in these conditions, tal model would also provide a platform for the
granulosa cells proliferated and formed cumu- investigation of epigenetic inheritance and trans-
lus–oocyte complexes. After culture in in vitro generational effects linked to environmental fac-
maturation conditions, 33 of 116 oocytes (28%) tors. The ability to generate large numbers of
extruded a first polar body, marking the comple- fully grown oocytes would facilitate biochemi-
tion of the first meiotic division. cal and proteomic analyses to complement the
In vitro fertilization of mature stem cell– imaging-based studies to which oocytes so beau-
derived oocytes was followed by development to tifully lend themselves. Accurate in vitro models
the two-cell stage, and 26% of these two-cell would also allow a gauge of precision of germ-
embryos progressed to the blastocyst stage. Of line gene-editing tools.
the 212 two-cell embryos that were transplant- If the in vitro derivation of human oocytes
ed into oviducts of pseudopregnant dams, 11 becomes feasible, the oocytes could be used to
(5%) resulted in live offspring. Hayashi and col- treat women with ovarian insufficiency caused
leagues had previously obtained similar results by genetic or environmental factors or by medi-
in fetal mouse gonadal cells, which suggested cal interventions such as gonadotoxic cancer
that their embryonic stem-cell–derived gonadal therapies, including gamma irradiation. Human
cells had similar properties to the authentic cells.5 oocytes could also be used to treat women who
However, there is merit in further research into have difficultly conceiving owing to advanced
the epigenetic program, the efficiency of meiotic reproductive age and to supply donor oocytes for
recombination and accuracy of chromosome seg- treatments such as mitochondrial-replacement
regation, and the ability to accumulate maternal therapy. However, any clinical use of embryonic
transcripts and organelles such as mitochondria stem-cell–derived oocytes would require very
during oocyte growth. Such research could poten- stringent safety tests and consideration of the
tially result in greater efficiencies in generating ethical and legal implications.
fertilizable oocytes that yield viable pregnancies. Disclosure forms provided by the authors are available at
Translating the work on mouse gonadal cells NEJM.org.

by Hayashi and colleagues to humans would be From the Newcastle Fertility Centre, Biosciences Institute,
challenged by gaps in our knowledge of the de- Newcastle University, and the Centre for Life, Newcastle upon
velopmental trajectory of human germ cells and Tyne (M.H.), and the Gurdon Institute, University of Cam-
bridge, Cambridge (A.S.) — all in the United Kingdom.
ovarian somatic cells. Moreover, germ-cell differ-
entiation and oogenesis occur over a very pro- 1. Yoshino T, Suzuki T, Nagamatsu G, et al. Generation of ovar-
tracted period in humans, which may be techni- ian follicles from mouse pluripotent stem cells. Science 2021;​
cally challenging to recapitulate in vitro. The 373(6552):​eabe0237.
2. McLaren A. Primordial germ cells in the mouse. Dev Biol
history of translating landmark developments 2003;​262:​1-15.
from mouse to human is notable for long inter- 3. Dokshin GA, Baltus AE, Eppig JJ, Page DC. Oocyte differen-
vals of time: the derivation of the first human tiation is genetically dissociable from meiosis in mice. Nat Genet
2013;​45:​877-83.
embryonic stem cells lagged behind that of 4. Cahoon CK, Hawley RS. Regulating the construction and
mouse embryonic stem cells by 17 years. Deriva- demolition of the synaptonemal complex. Nat Struct Mol Biol
tion of functional human oocytes in vitro is likely 2016;​23:​369-77.
5. Hikabe O, Hamazaki N, Nagamatsu G, et al. Reconstitution
to require patience and persistence. Nonethe- in vitro of the entire cycle of the mouse female germ line. Nature
less, this objective is worth pursuing because of 2016;​539:​299-303.
the potential benefits to infertile women and the DOI: 10.1056/NEJMcibr2112049
rescue of endangered mammals. Copyright © 2022 Massachusetts Medical Society.

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