Gonadotrope remodeling in sustained low estrogen states: single-cell transcriptomic analysis reveals gonadotrope subtypes and activation of stem cell populations†.

To study the effect of low-estrogen and the menopause state upon female pituitary function, we used the 4-vinylcyclohexene diepoxide (VCD)-induced ovarian failure model to gradually reduce serum estradiol (E2) levels. We utilized single-cell RNA sequencing transcriptomics analysis (scRNA-seq) to determine how reduced E2 levels influence pituitary gonadotrope gene expression and cell state, with a focus on potential contributions from pituitary stem cells to gonadotrope population growth. VCD-treated mice were acyclic and exhibited 8-15-fold increases in serum levels of the gonadotropins follicle-stimulating hormone and luteinizing hormone. Serum E2 levels were half those of normally cycling proestrous females. Pituitary scRNA-seq identified two gonadotrope populations based upon distinct gene expression signatures (a higher cell number primary cluster, designated here as GON1 and a smaller cell number secondary cluster, designated GON2). The GON1 population increased in number significantly in low-E2, VCD-treated mice, expressing markers indicative of contributions from pituitary stem cell activation. Both gonadotrope populations showed upregulated genes for canonical pathways supporting mRNA translation and hormone secretion, which correlated well with the high serum gonadotropin levels. Consistent with the high serum gonadotropin levels, Fshb and Lhb mRNAs were increased in whole pituitary samples (quantitative real time polymerase chain reaction). The pituitary Sox2-positive stem cell population exhibited increased expression of more mature transitional progenitors, including Sox9 mRNA, and showed upregulation of pathways involved in stem cell development and mitotic activity. This model provides insight into the cellular and molecular adaptations of the pituitary to the low-E2, menopausal state, with potential relevance for understanding human perimenopausal and menopausal physiology.