Original article
Wnt signalling in testicular descent: A candidate mechanism for cryptorchidism in Robinow syndrome,☆☆

Presented at the 2011 American Academy of Paediatrics National Conference and Exhibition, Boston, USA.
https://doi.org/10.1016/j.jpedsurg.2012.08.038Get rights and content

Abstract

Background/Aims

Robinow syndrome is caused by mutations in Wnt-5a or its receptor Ror2 and can lead to cryptorchidism, though the mechanisms are unclear. Wnt-5a knock-out mice fail to undergo gubernacular swelling, similar to insulin-like hormone 3 (INSl-3) knock-out mice. We aimed to characterise Wnt-5a and Ror2 expression in rat gubernacula to better understand how Wnt-5a signalling affects testicular descent.

Methods

Sprague–Dawley rats (n = 27) were collected with ethics approval (A644) at embryonic days (E) 15, 17, 19 and postnatal day (D) 2. Control and antiandrogen-treated groups were processed for immunohistochemistry for Wnt-5a, Ror2 and β-catenin. Sagittal sections were examined using confocal microscopy.

Results

Wnt-5a and Ror2 were strongly expressed in the gubernacular bulb at E17 controls, their levels declining at E19 and almost absent by D2. Wnt-5a significantly co-localised with the important transcription factor β-catenin at E17. There was no obvious difference in staining with androgen blockade.

Conclusion

Wnt-5a, through Ror2 and β-catenin may play a vital role in regulating the gubernacular swelling reaction downstream of INSL-3. Human mutations in Wnt-5a or Ror2 could prevent early gubernacular growth, as suggested by undescended testes in 70% of patients with Robinow Syndrome.

Section snippets

Methods

All experiments were approved by the institutional animal ethics committee (A644). Time-mated Sprague–Dawley dams were randomly allocated into either a control group or a group administered the anti-androgen flutamide according to a previously described method [11]. Pregnant dams were kept in the institute's Animal Research Laboratory and housed in standard shoe-box cages in a temperature-controlled environment with 12-h light/dark cycles. Animals were fed commercial rat chow and had access to

Wnt-5a was expressed in the E17 rat gubernaculum (n = 10)

At E17, Wnt-5a was strongly expressed in the tip of the male rat gubernaculum and around cells of the developing cremaster muscle. Expression appeared reduced in both areas by E19 (n = 10) and was absent by D2 (n = 9), coinciding with the beginning of inguino-scrotal migration (Fig. 1A-C). There was no qualitative difference in staining between control and antiandrogen-treated rats at E19 ( and D2 (n = 7) (not shown).

The Wnt-5a receptor Ror2 was maximally expressed in the E17 gubernaculum (n = 10 gubernacula)

Ror2, the non-canonical receptor for Wnt-5a, was strongly expressed in the

Discussion

We report for the first time, immunolocalisation of Wnt-5a and its receptor Ror2 within the rat gubernaculum. Wnt-5a expression was strongest in the gubernacular bulb and cremaster muscle at E17 during the critical period of active growth and swelling known to be controlled by INSL-3, and required to facilitate transabdominal testicular descent [19], [20]. Levels of Wnt-5a within the bulb rapidly decreased following gubernacular eversion and migration towards the scrotum (D2). Similarly,

Limitations

Although fluorescence immunohistochemistry is useful for visualisation of particular antigens, quantification of antigen expression is sometimes difficult to ascertain. It was not possible to conduct rigorous statistical analyses which could confirm the observed changes of these factors over time. However, it was possible to replicate the qualitative immunohistochemical findings in many specimens. Alternate techniques such as real-time PCR would be more suitable for the purposes of

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    Supported in part by the National Health and Medical Research Council (Australia), Grant no. ID60736.

    ☆☆

    Presented at the 2011 American Academy of Pediatrics National Conference and Exhibition, Boston, USA.

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