Abstract
In vitro folliculogenesis of primordial and early preantral follicles is necessary for increment of reproductive efficiency in domestic animals, humans and endangered species. Recent study in phosphatase and tensin homolog (Pten) -knockout mice has revealed that this phosphatase acts as an inhibitory factor in follicle activation of primordial pool with the resultant inhibition of oocyte growth. To test in vitro effect of a phosphatase inhibitor on growth initiation of isolated non-growing oocytes in neonatal ovaries, we applied a specific inhibitor (bpV (HOpic)) for PTEN in culturing system. Non-growing oocytes isolated from the ovaries of newborn BDF1 (C57BL/6 × DBA/2) pups were divided to four culture groups. Five days after culture, the oocytes in 14 μmol/l bpV only, 14 μmol/l bpV plus 100 ng/ml Kit Ligand (KL), and 100 ng/ml KL groups showed significantly (P<0.05) growth (19.3±0.55, 25.8±0.53 and 21.6±0.29 μm, respectively) compared with that of the control (no additive) (16.9±0.53 μm). In addition, western blotting in those groups showed enhanced expression of phosphorylated Akt. In conclusion, we clearly demonstrate that isolated non-growing oocytes develop in phosphatase inhibitor, especially to PTEN, incorporated culturing system, and show first as we know that oocytes with zona Pellucidae can be obtained in vitro from isolated non-growing oocytes.
Highlights
In the mammalian ovaries, several to ten thousands of primordial follicle’s oocytes are contained at birth, and these oocytes situate in the ovarian cortex in long duration of the fertile period [1,2,3]
Recent study in phosphatase and tensin homolog (Pten) -knockout mice [10] has revealed that this phosphatase acts as an inhibitory factor in follicle activation of primordial follicle pool
We show first as we know that phosphatase inhibitor, especially to PTEN can stimulate in vitro growth activation of isolated non-growing primary oocytes and that more effectively concomitant with Kit Ligand (KL) the oocytes can be grown with formation of their surrounding zona pellucidae
Summary
Several to ten thousands of primordial follicle’s oocytes (non-growing primary oocytes) are contained at birth, and these oocytes situate in the ovarian cortex in long duration of the fertile period [1,2,3]. Primordial follicles start growing with “follicle growth initiation or follicle activation” from primordial follicle pool, substantially evaluated by growth initiation of primary oocytes included, which is termed as the entry of primordial follicles into primary follicle stage [1,3]. This activation is indispensable for oocyte growth. Considered together that the study of PTEN knockout mouse showed precocious follicle development of all primordial follicles, it positively motivated us to develop a method feasible for more efficient growth initiation of primary oocytes by incorporation of PTEN inhibitor in culture media
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