Expression of constitutively active Notch1 in male genital tracts results in ectopic growth and blockage of efferent ducts, epididymal hyperplasia and sterility

Abstract

The Notch signaling pathway is involved in a variety of developmental processes. Here, we characterize the phenotypes developing in the reproductive organs of male transgenic (Tg) mice constitutively expressing the activated mouse Notch1 intracellular domain (Notch1(intra)) under the regulatory control of the mouse mammary tumor virus (MMTV) long terminal repeat (LTR). Tg expression was detected in testis, vas deferens and epididymis by Northern blot analysis. In situ hybridization with a Notch1-specific probe lacked sensitivity to detect expression in normal-appearing cells, but demonstrated expression in hyperplastic epithelial cells of the vas deferens, epididymis and efferent ducts. Tg males from three independent founder lines were sterile. Histological analysis of reproductive organs of young Tg males (postnatal ages 8 and 21) showed no difference compared to those of non-Tg males. In contrast, in adult Tg mice from day 38 onwards, the efferent ducts, the vas deferens and most epididymal segments revealed bilateral epithelial cell hyperplasia with absence of fully differentiated epithelial cells. Electron microscopy confirmed the uniformly undifferentiated state of these cells. Immunohistochemistry with anti-PCNA antibody also revealed enhanced proliferation of Tg epididymis. In adult Tg testis, the different generations of germ cells of seminiferous tubules appeared normal, although some tubules were highly dilated and revealed an absence of early and/or late spermatids. The epithelial cells of the Tg tubuli recti and rete testis were not abnormal, but the rete testis was highly dilated and contained numerous spermatozoa, suggesting a downstream blockage. Consistent with a blockage of efferent ducts often seen at the rete testis/efferent duct interface, spermatozoa were absent in epididymis of all adult Tg mice and in all highly hyperplastic efferent duct tubules of these Tg mice. Such a blockage was visualized by injection of Evans blue dye into the rete testis lumen. Finally, the presence of ectopic hyperplastic efferent duct tubules was observed within the testicular parenchyma itself, outside their normal territory, suggesting that Notch1 signaling is involved in the establishment of these borders. This phenotype seems to represent a novel developmental defect in mammals. Together, these results show that constitutive Notch1 signaling significantly affects the development of male reproductive organs.

PTEN signaling through RAF1 proto-oncogene serine/threonine kinase (RAF1)/ERK in the epididymis is essential for male fertility

“…The most pronounced change in cellular organization in initial-segmentspecific Pten knockouts was extensive epithelial folding. In the male reproductive tract, epithelial folding has only been observed in mouse efferent ducts after constitutive activation of NOTCH1 signaling (18) and has not been observed in other Pten knockout models. During normal tissue morphogenesis, epithelial folding is mediated by cell shape changes driven by apical constriction or, alternatively, by differential positioning of adherens junctions (19).…”

Activation of Notch-Mediated Protective Signaling in the Myocardium

“…Activated Notch overexpression stimulates proliferative signaling in the mouse male genital tract 45 and in cardiomyocytes infected with adNICDEGP ( Figure 6). Previous studies have investigated the role of embryonic signaling in myocardial repair.…”

“…45 PCNA is clearly detected in the nucleus of adNICDEGFP-overexpressing myocytes ( Figure 6A), but nuclear localization is not seen in adEGFP-infected cells ( Figure 6B), whereas surviving myocytes expressing adenoviral activated Notch stained positive for Ki67 ( Figure 6C). Collectively, these results suggest activated Notch stimulates proliferative signaling in cardiomyocytes.…”

Why are epididymal tumours so rare?

“…9 Notch1 mRNA is in all epididymal regions, without cellular identity; 10 Notch1 is only detectable by in situ hybridisation in hyperplastic epididymal epithelial cells of constitutively active Notch1-transgenic mice. 11 These embryonic stem cell factors may have other functions, such as cellular differentiation, in the adult epididymis.…”