Abstract
BackgroundRecent findings have revealed that the female gonad may have regenerative activity with having germ line stem cells in juveniles and adults. Application of these germ line stem cells could be an alternative therapy for reproductive disorders in regenerative medicine.MethodsTo enhance the potency of differentiation into oocyte-like cells (OLCs) and folliculogenesis, we overexpressed Oct4 in ovarian stem/stromal cell (OvSCs) and examined the cellular properties related to stemness and self-renewal ability and finally demonstrated the ability of in vitro differentiation and folliculogenesis.ResultsOvarian cortex included putative stem cells in terms of AP activity, cell cycle status, cell proliferation, expression of mesenchymal lineage surface markers and pluripotent transcriptional markers. Further, Oct4 transfected OvSCs (Oct4-OvSCs) were enhanced their AP activity and cell proliferation compared to OvSCs. The potential on in vitro differentiation into OLCs and in vivo folliculogenesis was also evaluated in OvSCs and Oct4-OvSCs, respectively. Oct4-OvSCs possessed higher oogenesis potential in vitro than OvSCs, in terms of expression of germ cell markers by RT-PCR and the number of OLCs. When OvSCs and Oct4-OvSCs were xeno-transplanted into infertile mice ovaries, the OvSCs transplantation induced new primary follicle formation and hormonal levels of estradiol and FSH remained similar to that of normal mice. However, Oct4-OvSCs possessed higher ability for folliculogenesis based on inducing developing follicles with thecal layer and granulosa cells and more similar estradiol level to normal mice.ConclusionsThese findings demonstrated that putative stem cells were present in ovarian cortex and exhibited differentiation ability into OLCs and folliculogenesis in vivo, and Oct4-overexpression enhanced these ability, suggesting their cellular models based on gene therapy in understanding the mechanisms of oogenesis and folliculogenesis, and finally in view of reproductive cell therapy.
Highlights
Recent findings have revealed that the female gonad may have regenerative activity with having germ line stem cells in juveniles and adults
Increased alkaline phosphatase (AP) activity in Oct4-ovarian stem/stromal cells (OvSCs) In primary culture, the OvSCs were grown as heterogeneous populations with elongated, sphere-like or spindlelike adherent features
OvSCs were transfected with lentiviral vector which expressed GFP as an infection control, and the infection rate in Oct4-OvSCs was found to be 66.7 ± 0.7 %, while non-transfected cells were not detected for GFP (Fig. 1e)
Summary
Recent findings have revealed that the female gonad may have regenerative activity with having germ line stem cells in juveniles and adults Application of these germ line stem cells could be an alternative therapy for reproductive disorders in regenerative medicine. The recent findings have shown that the female gonad of juveniles and adults may have regenerative activity and possess germ line stem cells, which can be considered as an alternative regenerative medicine therapy for reproductive disorders. Similar to mesenchymal stem cells (MSCs), which shows ability for self-renewal and differentiation into multiple mesenchymal lineages in vitro [10], OvSCs known to express Oct, Sox and Nanog, which are considered to be an important early transcription factors responsible for the regulation of stem cells pluripotency, and revealed the expression of mesenchymal lineage-cell surface markers such as CD44, CD29 and CD90 [11]. The evaluation of germ cell specific markers [growth differentiation factor 9b (GDF9b), C-Mos, deleted in azoospermia-like (DAZL), Vasa, zona pellucida C (ZPC), Stella and c-kit] and folliculogenesis marker [follicular stimulating hormone receptor (FSHR)] expression is an important criteria for characterizing OLCs in addition to the assessment of pluripotent transcriptional marker (Oct4) expression [12,13,14]
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