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Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice

  • Authors:
    • Qinyue Cao
    • Hehua Wang
    • Jingjing Hu
    • Yan Wang
    • Tong Dai
    • Fen Liu
    • Xia Yang
    • Qinyu Yang
    • Chunhua Tu
  • View Affiliations / Copyright

    Affiliations: Department of Obstetrics and Gynecology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China
    Copyright: © Cao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 265
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    Published online on: July 22, 2025
       https://doi.org/10.3892/mmr.2025.13630
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Abstract

Cellular prion protein (PrPc) regulates ovarian reserve maintenance through anti‑Müllerian hormone (AMH)‑dependent mechanisms. The present study explored the role of PrPc in the ovarian function of mice using complementary in vitro and in vivo models. First, prion protein gene (PRNP) knockdown or overexpression was carried out in mouse ovarian granulosa cells. In vitro analyses conducted using flow cytometry and ELISA revealed that the depletion of PrPc specifically hindered the secretion of AMH compared with control groups, while the levels of progesterone (P4) and estradiol (E2) remained unchanged across all experimental groups. Importantly, the reduction in AMH levels was reversed upon re‑expression of PrPc. Additionally, neither the distribution of the cell cycle nor the rates of apoptosis were affected by the manipulation of PrPc. Subsequently, a comparative analysis of mice with PRNP knockout (KO) vs. wild‑type mice was performed. However, PrPc depletion did not alter the production of progesterone or estradiol. Whilst the ovarian histology remained intact in KO mice, an elevation in follicle‑stimulating hormone levels was observed, thereby suggesting a potential involvement of compensatory neuroendocrine regulation. These findings revealed that PrPc may be a novel modulator for maintaining the ovarian reserve which depends on AMH. The present study redefined the molecular landscape of ovarian reserve depletion by identifying the dysfunction of the PrPc‑AMH axis as a possible reason for diminished ovarian reserve syndromes.
View Figures

Figure 1

PrPc knockdown and OE
efficiency in mGCs. (A) mRNA expression levels of PRNP and
(B) protein expression levels of PrPc in mGCs
transfected with different shRNAs. (C) mRNA expression levels of
PRNP and (D) protein expression levels of PrPc in
mGCs transfected with OE vectors. β-actin was used as an internal
control. *P<0.05; **P<0.01; ***P<0.001. ns,
non-significant; PrPc, cellular prion protein;
PRNP, prion protein gene; mGCs, mouse ovarian granulosa
cells; OE, overexpression; sh, short hairpin; NC, negative
control.

Figure 2

Cellular prion protein exhibits
minimal influence on the cell cycle distribution and apoptosis of
mouse granulosa cells. Flow cytometry was employed to analyze the
cell cycle distribution, enabling quantification of cells in the
G0/G1, S and G2/M phases after
transfection with (A) different shRNAs or (B) OE vectors. Apoptotic
and necrotic cells were assessed using Annexin V/PI staining after
transfection with (C) different shRNAs or (D) OE vectors. ns,
non-significant; sh, short hairpin; NC, negative control; OE,
overexpression.

Figure 3

Cellular prion protein knockdown
specifically reduces AMH secretion in mGCs. (A) AMH, (B) P4 and (C)
E2 levels in the culture medium of mGCs transfected with
different shRNAs, OE vectors or their combination. *P<0.05. ns,
non-significant; AMH, anti-Müllerian hormone; mGCs, mouse granulosa
cells; P4, progesterone; E2, estradiol; sh, short
hairpin; OE, overexpression; NC, negative control.

Figure 4

Prion protein gene KO mice exhibit
normal ovarian morphology and follicular development. (A)
PrPc protein expression in the ovaries of WT and KO
mice. (B) Ratio of bilateral ovary weight to body weight showed no
differences between groups. (C) H&E staining on mouse ovaries
(scale bars, 200 µm for overviews and 100 µm for higher mag). The
stages of follicular development are clearly indicated by
color-coded arrows: Blue represents primordial follicles, which
consist of a single layer of squamous granulosa cells; green
denotes primary follicles, characterized by cuboidal granulosa
cells; white signifies secondary follicles, identifiable by having
≥3 layers of granulosa cells and the early formation of an antrum;
black marks antral follicles, distinguished by the presence of a
fluid-filled cavity; and yellow indicates atretic follicles, which
display pyknotic granulosa cell nuclei and fragmentation of the
zona pellucida. (D) Follicle counts across developmental stages
were comparable between WT and KO mice. ns, non-significant; WT,
wild-type; KO, knockout; PrPc, cellular prion
protein.

Figure 5

KO mice exhibit elevated FSH levels
but normal E2, AMH and LH levels. Serum hormone levels
of (A) FSH, (B) E2, (C) AMH and (D) LH in WT and KO
mice. **P<0.01. ns, non-significant; FSH, follicle-stimulating
hormone; LH, luteinizing hormone; E2, estradiol; AMH,
anti-Müllerian hormone; WT, wild-type; KO, knockout.

Figure 6

KO mice exhibit regular estrous cycle
and fertility. (A) Vaginal secretion smears stained with H&E
stain during estrous cycle phases: Proestrus, estrus, metestrus,
diestrus (scale bar, 50 µm). (B) Regular estrous cycle patterns in
mice spanning 9 consecutive days. (C) Interval from mating,
following co-housing, to parturition. (D) Number of offspring at
birth. ns, non-significant; WT, wild-type; KO, knockout; P,
proestrus; E, estrus; M, metestrus; D, diestrus.
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Copy and paste a formatted citation
Spandidos Publications style
Cao Q, Wang H, Hu J, Wang Y, Dai T, Liu F, Yang X, Yang Q and Tu C: Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice. Mol Med Rep 32: 265, 2025.
APA
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F. ... Tu, C. (2025). Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice. Molecular Medicine Reports, 32, 265. https://doi.org/10.3892/mmr.2025.13630
MLA
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F., Yang, X., Yang, Q., Tu, C."Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice". Molecular Medicine Reports 32.4 (2025): 265.
Chicago
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F., Yang, X., Yang, Q., Tu, C."Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice". Molecular Medicine Reports 32, no. 4 (2025): 265. https://doi.org/10.3892/mmr.2025.13630
Copy and paste a formatted citation
x
Spandidos Publications style
Cao Q, Wang H, Hu J, Wang Y, Dai T, Liu F, Yang X, Yang Q and Tu C: Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice. Mol Med Rep 32: 265, 2025.
APA
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F. ... Tu, C. (2025). Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice. Molecular Medicine Reports, 32, 265. https://doi.org/10.3892/mmr.2025.13630
MLA
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F., Yang, X., Yang, Q., Tu, C."Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice". Molecular Medicine Reports 32.4 (2025): 265.
Chicago
Cao, Q., Wang, H., Hu, J., Wang, Y., Dai, T., Liu, F., Yang, X., Yang, Q., Tu, C."Role of cellular prion protein in mouse granulosa cells and its effects on ovarian function in knockout mice". Molecular Medicine Reports 32, no. 4 (2025): 265. https://doi.org/10.3892/mmr.2025.13630
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