Ovarian preantral follicles are responsive to FSH as early as the primary stage of development

in Journal of Endocrinology
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  • 1 Department of Animal Science, University of California Davis, Davis, California, USA
  • 2 Department of Applied Mathematics and Computer Science, Technical University of Denmark, Kongens Lyngby, Denmark

Correspondence should be addressed to A C Denicol: acdenicol@ucdavis.edu
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Follicle-stimulating hormone (FSH) is required for ovarian antral folliculogenesis and steroidogenesis, and there is increasing evidence that it may play critical roles in preantral follicle development. We hypothesized that preantral follicles begin responding to FSH as early as the primary stage of development. Our objectives were to establish whether the FSH receptor (FSHR) was expressed in bovine preantral follicles and to determine the effects of FSH in these follicles and the surrounding ovarian tissue. Preantral follicles were isolated from bovine ovaries and subjected to immunolocalization of FSHR. Ovarian cortical strips were cultured with FSH or vehicle for 2 or 4 days and subjected to RNA sequencing, hematoxylin/eosin staining and immunostaining for p42/44 MAPK. Finally, cortical strips were cultured for 4 days with FSH before Western blot analysis of total and phosphorylated p42/44 MAPK and total aromatase. We found greater FSHR labeling intensity per cell in preantral follicles at the primary stage compared to other stages (P < 0.05). FSH upregulated genes involved in energy metabolism and MAPK signaling and downregulated genes related to phagosome and allograft rejection in the ovarian cortex. Preantral follicles cultured in situ with FSH had greater expression of total p42/44 MAPK (P < 0.05), but no difference was detected in whole tissue Western blot for phosphorylated p42/44 MAPK or aromatase. We conclude that the FSHR is expressed in preantral follicles as early as the primary stage of development, and that FSH upregulates cell metabolism and activates MAPK signaling pathways in preantral follicles.

Supplementary Materials

    • Supplementary Table 1. Counts per million (CPM) of each of the 235 transcripts identified in the time course analysis and clustered into clusters 1-5. Rep = replicate; D2 = day 2 of culture; D4 = day 4 of culture; FC = fold change. Due to log transformation of the raw expression data, fold changes relative to day 0 (D0) are 2subtracted CPM.
    • Supplementary Figure 1. Negative controls for immunofluorescent labeling. (A) Bovine COC labeled with rabbit isotype IgG control; (B) Isolated preantral follicle no-primary control, (C) Antral follicle cryosection no-primary control; arrows pointing to granulosa cells; (D) Ovarian cortex cryosection with preantral follicle (denoted by arrowhead) rabbit isotype IgG control. TC: theca cells; AC: antral cavity. Scale bars = 50 µm for A, B, and C. Scale bar = 100 µm for D.
    • Supplementary Figure 2. Regression analysis of intensity of FSHR staining according to follicle stage and cell number.

 

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