Dysregulation of Notch-FGF signaling axis in germ cells results in cystic dilation of the rete testis in mice

Abstract

Numb (Nb) and Numb-like (Nbl) are functionally redundant adaptor proteins that critically regulate cell fate and morphogenesis in a variety of organs. We selectively deleted Nb and Nbl in testicular germ cells by breeding Nb/Nbl floxed mice with a transgenic mouse line Tex101-Cre. The mutant mice developed unilateral or bilateral cystic dilation in the rete testis (RT). Dye trace indicated partial blockages in the testicular hilum. Morphological and immunohistochemical evaluations revealed that the lining epithelium of the cysts possessed similar characteristics of RT epithelium, suggesting that the cyst originated from dilation of the RT lumen. Spermatogenesis and the efferent ducts were unaffected. In comparisons of isolated germ cells from mutants to control mice, the Notch activity considerably increased and the expression of Notch target gene Hey1 significantly elevated. Further studies identified that germ cell Fgf4 expression negatively correlated the Notch activity and demonstrated that blockade of FGF receptors mediated FGF4 signaling induced enlargement of the RT lumen in vitro. The crucial role of the FGF4 signaling in modulation of RT development was verified by the selective germ cell Fgf4 ablation, which displayed a phenotype similar to that of germ cell Nb/Nbl null mutant males. These findings indicate that aberrant over-activation of the Notch signaling in germ cells due to Nb/Nbl abrogation impairs the RT development, which is through the suppressing germ cell Fgf4 expression. The present study uncovers the presence of a lumicrine signal pathway in which secreted/diffusible protein FGF4 produced by germ cells is essential for normal RT development.

Rhox8 homeobox gene ablation leads to rete testis abnormality and male subfertility in mice

“…Both WT and Rhox8 KO showed normal Sertoli cells of valve-like structure (Figure 5C). A previous study revealed that dysregulation of Notch-FGF signaling contributes to a dilated RT in mice [21]. Hence, we investigated the expression of Notch target genes, Hes1 and Hey1 , as well as Fgf4 in the testes.…”

“…The relative levels of mRNA were calculated using the 2 -Ct method. The primers for gene amplification have been previously reported [10,13,[17][18][19][20][21] or are listed in Supplementary Table S1.…”

Aberrant activation of KRAS in mouse theca-interstitial cells results in female infertility

“…Immunohistochemistry was performed by an avidin-biotin immunoperoxidase method as described previously (Cao et al, 2022). Briefly, deparaffinized sections were rehydrated and then incubated with 1% H 2 O 2 for 30 min.…”

“…Western blotting was used to determine the effectiveness of Foxo1 siRNAs in suppressing FOXO1 protein levels in theca cells. A transient DNA transfection procedure described previously was used to overexpress mouse Foxo1 in tKrasMT theca cells (Cao et al, 2022). Briefly, the cells were cultured to approximately 80% confluence in 12-well plates and were transiently transfected with 1 µg of pCMV-Foxo1 plasmid DNA (Nakae et al, 2001) (Addgene, Watertown, MA, USA) with 2 µL of X-TremeGene 360 reagent (Roche, Mannheim, Germany) per well for 72 h. The cells were then cultured in RPMI-1640 containing 10% FBS.…”

A comparative analysis of genes differentially expressed between rete testis cells and Sertoli cells of the mouse testis

“…In addition, SCs in the transition region exhibit an immature phenotype and prolonged proliferation in hamsters 3 and rats 4 , 5 which also might be influenced by the RT. It has been reported that seminiferous tubules also may affect RT function by luminal factors 6 .…”