Identification of follicle-stimulating hormone-responsive genes in Sertoli cells during early postnatal mouse testis development

Abstract

Background: In the mouse testis, Sertoli cells rapidly divide during a narrow window of time pre-pubertally and differentiate thereafter. The number of Sertoli cells determines the testis size and germ cell-carrying capacity. Follicle-stimulating hormone (FSH) binds its cognate FSH-receptors expressed on Sertoli cells and acts as a mitogen to regulate their proliferation. Fshb^-/- mutant adult male mice have reduced Sertoli cell number and testis size and reduced sperm number and motility. However, FSH-responsive genes in early postnatal mouse Sertoli cells are unknown.

Objectives: To identify FSH-responsive genes in early postnatal mouse Sertoli cells.

Materials and methods: A fluorescence-activated cell sorting method was developed to rapidly purify Sertoli cells from control and Fshb^-/- mice carrying a Sox9 ^GfpKI allele. These pure Sertoli cells were used for large-scale gene expression analyses.

Results: We show that mouse Sertoli cells rarely divide beyond postnatal day 7. Our in vivo BrdU labeling studies indicate loss of FSH results in a 30% reduction in Sertoli cell proliferation in mice at 5 days of age. Flowsorted GFP⁺ Sertoli cells with maximal Fshr expression were 97%-98% pure and mostly devoid of Leydig and germ cells as assessed by Taqman qPCR quantification of gene expression and immunolabeling of the corresponding cell-specific markers. Large-scale gene expression analysis identified several differentially regulated genes in flow-sorted GFP⁺ Sertoli cells obtained from testis of control and Fshb^-/- mice at 5 days of age. The top 25 networks identified by pathway analysis include those related to the cell cycle, cell survival and most importantly, carbohydrate and lipid metabolism and molecular transport.

Discussion: Several of the FSH-responsive genes identified in this study could serve as useful markers for Sertoli cell proliferation in normal physiology, toxicant-induced Sertoli cell/testis injury, and other pathological conditions.

Conclusion: Our studies reveal that FSH-regulates macromolecular metabolism and molecular transport networks of genes in early postnatal Sertoli cells most likely in preparation for establishment of functional associations with germ cells to successfully coordinate spermatogenesis.

Keywords: FSH; FSHR; proliferation; sertoli cells; spermatogenesis; testis.