Review Article

Stem Cell Interaction with Somatic Niche May Hold the Key to Fertility Restoration in Cancer Patients

Table 1

Consolidation of published literature on stem cells in adult mammalian ovary based on the concept of pluripotent (VSELs) and progenitor stem cell population (OGSCs).

ReferenceStudy highlightsInterpretation of published literature

Johnson et al. [7]
Germline stem cells and follicular renewal in the postnatal mammalian ovary
Mitotically active SCP3+ and MVH+ germline stem cells in surface epithelium of adult mice ovary
Chimeric follicles observed when wild type ovarian tissue is grafted onto ovary of GFP expressing transgenic mice
Several groups including our results also report the presence of stem cells in the ovary surface epithelium
Probably they detected the bigger OGSCs since the cells were SCP3+ and MVH+
Chimeric follicles suggest that the oocyte and granulosa cells do not originate from a common bipotent progenitor stem cell as suggested by Bukovsky et al. [26, 61, 62]

Johnson et al. [63]
Oocyte generation in adult mammalian ovaries by putative germ cells derived from bone marrow and peripheral blood
Extraovarian bone marrow (BM) origin of germ stem cells (GSCs) in adult mice and distribution by peripheral blood (PB) to the ovaries
BM and PB express primordial germ cell markers Oct-4, Dazl, Mvh, Stella, Fragilis, and Nobox
Compromised ovaries in young mice (due to chemotherapy) possibly mobilized VSELs from the bone marrow to enter circulation
In addition to the pluripotent markers (Oct-4 and Nanog), the mobilized VSELs expressed germ cell specific markers. On similar note during stroke the mobilized VSELs exhibit neural markers like GFAP, nestin, beta-III-tubulin, Olig1, Olig2, Sox-2, and Musashi [35]
Possibly the VSELs sense the nature of damage—and thus proliferate and give rise to progenitor stem cells exhibiting specific markers
Interestingly the mobilized cells reported were Lin- and Sca-, implying that they neither were of hematopoietic origin nor were pluripotent. The authors probably detected GSCs which expressed germ cell specific markers

Lee et al. [47]
Bone marrow transplantation generates immature oocytes and rescues long-term fertility in a preclinical mouse model of chemotherapy induced premature ovarian failure
Chemotherapy sterilized mice were transplanted BM cells from coat color mismatched donors
All pups born were of recipient germ line
BMT possibly provides an endocrine/paracrine signal that improves the functionality of ovarian niche there by restoring function
BM does not serve as a source of germ cells since all the pups are similar to the recipient

Bukovsky et al. [64]
Bone-marrow-derived cells and alternative pathways of oogenesis in adult rodents
Suggested alternative pathway of oogenesis in adult rodents
Explained that the rodent germ cells may, but do not necessarily originate from the OSE stem cells.
Proposed alternative origin of putative germ cells from the medullary region
Used neonatally estrogenized female rats which lack OSE but with normal stock of primordial follicles as study model
Showed clusters of SSEA1+ cells in the ovarian medulla-precursors of oocytes
Proposed that female germ cells should receive an impulse from the immune system-related cells to become oocytes. Therefore, if triggered by BM derived cells, the germ cells in ovarian medulla may represent an alternative source of oocytes for renewal of primary follicles
We propose that there may not be any alternative pathway existing in rat ovaries
Ovary after neonatal exposure to estradiol is compromised and its homeostasis is disturbed. This may mobilize VSELs from BM through PB. Mobilized VSELs possibly enter ovarian medulla through the blood vessels and then try to reach the cortex for follicular assembly
Their results probably show that SSEA1+ cells migrate from the BM into the medulla of the ovaries

Szotek et al. [65]
Normal ovarian surface epithelial label-retaining cells exhibit stem/progenitor cell characteristics
Identified a label-retaining cell (LRCs) population in coelomic epithelium of adult H2B-GFP transgenic mouse ovary
These cells exhibit quiescence, functional response to the estrus cycle, slow cycling, and may undergo asymmetric cell division, exhibit cytoprotective mechanisms by enrichment for side population, and show increased growth potential in vitro
The LRCs reported by them are possibly the VSELs which undergo asymmetric cell division
Interestingly VSELs do not stain with DAPI possibly because they mostly comprise of euchromatin whereas DAPI binds preferentially to heterochromatin [30]

Zhang et al. [66]
Expression of stem and germ cell markers within nonfollicle structures in adult mouse ovary
Used Oct-4-EGFP transgenic mouse model to study the expression of stem and germ cell markers in adult murine ovaries
OCT3/4, MVH, SSEA-1, and SCF-R in specific cell aggregates of 50–200 cells (distinct from follicles) within the adult mouse ovary
Aggregates have large round nuclei, intensely stain with Haematoxylin, positive for OCT-4, SSEA-1, SCF-R, and MVH; also have SCP3 and DMC1 (meiotic markers by RT-PCR); interestingly they lacked GDF-9 (a postmeiotic marker)
Authors conclude a mixed population of committed stem cells as well as transitional stage germline cells that might retain the capacity of proliferation and differentiation
These aggregates possibly represent clonal expansion of OGSCs with cytoplasmic continuity described as germ cell nests in developing fetal ovary [67]. We have observed similar structures in adult mouse and human ovary (Figure 1).
Like the OGSCs, cells comprising the germ cell nests have characteristic dark stained nuclei after H staining.
OGSCs are immediate progenitors of VSELs and since this involves a shift from euchromatin to a committed genome of a germ cell—extensive chromatin compaction, remodeling occurs—giving dark appearance after H stain. OGSCs divide rapidly to form germ cell nests.
This data directly supports postnatal oogenesis in adult mammalian ovary. However, the group have reported OGSCs and not VSELs

Zou et al. [22]
Production of offspring from a germline stem cell line derived from neonatal ovaries
Proliferative MVH positive (10–12 μm) large FGSCs purified from neonatal and adult mouse ovaries and maintained in vitro for months
These cells, after transplantation into ovaries of chemotherapy sterilized recipients, generate chimeric follicles that were fertilized and produced viable offspring
Such large VASA positive cells have been reported also by Zhang et al. [66]
However, they immunosorted the initial cells for establishing the cultures based on MVH, a germ cell marker and not an early stem cell marker

Zou et al. [23]
Improved efficiency of female germline stem cell purification using fragilis-based magnetic bead sorting
Use of Fragilis, an early germ cell marker, to enrich cells (10–12 μm) for initiating cultures—further enhanced isolation efficiency of mouse FGSCsPossibly sorted OGSCs based on the size of the cells sorted by them

Pacchiarotti et al. [24]
Differentiation potential of germ line stem cells derived from the postnatal mouse ovary
Demonstrated the presence of GSCs in adult mouse ovary using Oct-4-EGFP transgenic mouse model
Detected three different types of GFP-OCT-4 positive cells based on size, namely, small (10–15 μm) sized in OSE; medium (20–30 μm) and big (50–60 μm) oocytes in the follicles by flow cytometry
Ploidy analysis results showed that 70% of these cells were tetraploid (possibly oocytes) and 30% were diploid (stem cells).
They further showed that CD133+ cells exist in the ovary but do not co-localize with GFP-OCT-4 suggesting that germ line stem cells in ovary are distinct from the circulating CD133+ cells
Flow Cytometry data shows that diploid stem cells exist in ovary and are further of two sizes in agreement with our data
However, their approach of using GFP-OCT-4 mice did not allow them to differentiate between cytoplasmic and nuclear OCT-4 since GFP will be expressed by both the stem cells as both the transcripts are under the control of common Oct-4 promoter.
CD133+ cells are possibly the VSELs but it is intriguing that they did not co-express OCT-4
Thus whether nuclear OCT-4 positive VSELs express GFP or not needs further investigation

Gong et al. [25]
Embryonic stem cell-like cells established by culture of adult ovarian cells in mice
Ovarian stromal cells (<40 μm) were subcultured on fibroblast monolayer and colony-forming cells were characterized
Detected pluripotent stem cells in adult mice ovary which could be expanded in culture
Two ES-like cell lines were established which expressed pluripotent markers and formed embryoid bodies and teratomas
The group was unable to provide information on the exact location on the pluripotent stem cells since they used all cells of size less than 40 μm to establish cultures. They mention stromal origin of stem cells but cells for initiating cultures were obtained by mincing whole ovary which will include the OSE also
Ovarian smears used to demonstrate the presence of Oct-4 positive cells and also RNA was extracted from the whole ovary for RT-PCR to show the pluripotent transcripts—thus OSE as a source of pluripotent stem cells is not ruled out in their study

Bukovsky et al. [26, 61, 62]
Immunohistochemical studies of the adult human ovary: possible contribution of immune and epithelial factors to folliculogenesis
Origin of germ cells and formation of new primary follicles in adult human ovaries
Oogenesis in cultures derived from adult human ovaries
Putative germ cells within the OSE of adult human ovary and originate from OSE stem cells which differentiate from mesenchymal progenitors in the ovarian tunica albuginea
Scraped OSE cells from adult human ovary in culture form large oocyte-like cells and follicle-like structures
Put forth the concept of bipotent progenitors capable of differentiating into oocytes and granulosa cells
The model of bipotent progenitors giving rise to germ and granulosa cells does not explain the chimeric follicles reported by other groups [22]
Our results are in agreement with theirs that stem cells in OSE can generate oocyte-like structures in vitro

Virant-Klun et al. [2729]
Putative stem cells with an embryonic character isolated from the ovarian surface epithelium of women with no naturally present follicles and oocytes
Parthenogenetic embryo-like structures in the human ovarian surface epithelium cell culture in postmenopausal women with no naturally present follicles and oocytes
Stem cells in aged mammalian ovaries
Small (diameter 2–4 μm) round putative stem cells also able to forming oocyte-like cells in vitro isolated from human OSE
These cells expressed mRNA for pluripotent markers like Oct-4, SSEA-4, Nanog, and Sox-2
After 20 days of culture formed oocyte-like cells expressing VASA, c-KIT and ZP2 transcripts
Accompanying bubble-like putative stem cells growing in close contact with oocytes possibly acting like granulosa cells supplying essential cellular machinery to the developing germ cells
Oocytes derived from these putative stem cells in vitro underwent parthenogenetic activation to form blastocyst-like structures
Investigators concluded that they had discovered small cells with pluripotent characteristics comparable to VSELs found in other adult human tissues and organs
Surface epithelial location of the stem-like cells in postmenopausal ovaries reported by them matches initial reports of the location of presumptive GSC (MVH-BrdU double-positive cells) in juvenile and young adult mouse ovaries [7]
The cells reported are probably the VSELs

Parte et al. [30]
Detection, characterization, and spontaneous differentiation in vitro of very small embryonic-like putative stem cells in adult mammalian ovary
Two distinct populations of putative stem cells detected in scraped OSE of adult mammalian ovary, namely, VSELs (1–3 μm) and progenitor stem cells (4–7 μm) termed OGSCs
VSELs express nuclear OCT-4 whereas the OGSCs show cytoplasmic OCT-4
Pluripotent markers Oct-4, Oct-4A, Nanog, Sox-2, TERT, and Stat-3 in human and sheep OSE
c-KIT, DAZL, GDF-9, VASA, and ZP4 expressing oocyte-like cells spontaneously differentiate in three weeks cultures
VSELs are the quiescent stem cell population that undergo asymmetric cell division whereas the OGSCs are the progenitors similar to Adark SSCs in testis, undergo extensive proliferation, and form aggregates just like cytoplasmic bridges in testis [20]
VSELs are totipotent to pluripotent in nature and give rise to OGSCs which further differentiate into oocyte-like structures, parthenotes, neuronal-like cells, and so forth
Observed close association of developing oocytes with mesenchymal cells in vitro formed by EMT of the OSE cells in initial cultures, similar to the results published recently [41]. We propose that granulosa-like cells are formed by EMT
Thus VSELs differentiate to give rise to oocytes whereas the epithelial cells undergo EMT to form supporting granulosa-like cells—thus resulting in primordial follicle assembly