BAPS Paper
Molecular signals governing cremaster muscle development: Clues for cryptorchidism

https://doi.org/10.1016/j.jpedsurg.2013.11.049Get rights and content

Abstract

Background/Aim

Cryptorchidism affects 2-4% of newborn boys. Testicular descent requires the gubernaculum to differentiate into cremaster muscle (CM) during androgen-mediated inguino-scrotal descent, but the cellular mechanisms regulating this remodeling remain elusive. β-Catenin, a marker of canonical Wnt signaling, promotes myogenic genes and cellular adhesion. We aimed to determine if androgen receptor (AR) blockade altered β-catenin and its downstream myogenic proteins within the CM.

Method

Gubernacula from male rats (n = 12) and rats treated with anti-androgen, flutamide (n = 12) at E19, D0, D2 were processed for immunohistochemistry. Antibodies against β-catenin, embryonic myosin, and myogenin were visualized by confocal microscopy.

Results

At E19, β-catenin immuno-reactivity (IR) localized to the CM membrane. By D2, cytoplasmic β-catenin-IR was noted with overall β-catenin-IR decreasing. Myogenic proteins resided primarily in cells containing β-catenin on their plasma membrane. Embryonic myosin-IR was high at E19 and then decreased by D2, while myogenin-IR increased. AR blockade increased cytoplasmic β-catenin at D2 and reduced levels of both myogenic proteins.

Conclusion

Myogenic proteins are present in CM cells containing β-catenin. AR blockade did not alter cellular adhesion via β-catenin. In contrast, blocking AR prevented β-catenin entering the nucleus and impaired CM myogenesis. Mutations in this pathway may result in idiopathic cryptorchidism.

Section snippets

Methods

Sprague-Dawley rats were purchased from a commercial supplier and housed in the institute's Animal Research Laboratory in standard shoebox cages. Animals were maintained in a temperature-controlled atmosphere with a 12-hour light-dark cycle and fed commercial rat chow and water ad libitum. Ethical approval was gained from the Institutional Ethics Committee (license number A644) and with correct care taken in accordance with the National Health and Medical Research Council animal ethics

β-Catenin localises to the developing CM on the cell surface

Labelling experiments showed that β-catenin immuno-reactivity (IR) was most prevalent in the CM with β-catenin-IR residing on the whole plasma membrane of CM cells in E19 (n = 4) and D0 specimens (n = 4). β-catenin-IR appeared cytoplasmic in D2 specimens that labelled ubiquitously throughout the entire CM (n = 4). Additionally, β-catenin-IR appeared to fragment by D2, a time when the gubernaculum is everting before migration into the scrotum (Fig. 1). The overall IR pattern in the time-points

Discussion

Emerging evidence suggests that the key effector of the canonical Wnt pathway, β-catenin, is essential for testicular descent [13] and that it interacts with AR to enable expression of downstream myogenic proteins. β-Catenin is highly conserved throughout evolution, and the intracellular cascade that regulates its translocation to either the nucleus or the plasma membrane hinges on the molecular machinery that stabilizes it in the cytoplasm. Ligands such as Wnt1, Wnt3a and Wnt5a, initiate the

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