Quantitative measurement of the androgen receptor in prepuces of boys with and without hypospadias

Abstract

WHAT'S KNOWN ON THE SUBJECT? AND WHAT DOES THE STUDY ADD?: Despite diverse anatomical and histological trials in humans and animal models, the aetiology of hypospadias remains unknown and currently there is no clear molecular explanation about the emergence of this disease; however, genetic, endocrine and environmental mechanisms have been suggested. The aim of the present study was to quantify and compare the androgen receptor (AR; mRNA and protein) levels in 40 prepuces of boys with and without hypospadias using quantitative real-time polymerase chain reaction, Western Blot and standardised, automated immunohistochemistry. AR mRNA (P = 0.013) and AR protein (P = 0.014) was significantly elevated in the prepuces of boys with hypospadias compared with boys without hypospadias. Altogether our data indicate that elevated AR mRNA and protein levels can be considered as a biochemical response of an AR signalling defect as an identified cause in boys with hypospadias. Additionally, nuclear staining intensity for AR-protein in specimens of boys with hypospadias was higher than in boys with phimosis. To address the role of the androgen receptor (AR) on mRNA and protein levels in prepuces of boys with and without hypospadias. Data from 40 foreskin specimens of consecutive circumcised boys (20 with vs 20 without hypospadias) were enrolled in this prospective study. After surgery, samples were fixed in formaldehyde and frozen in liquid nitrogen. Total RNA was isolated from frozen tissue and transcribed to complementary DNA. The amount of AR mRNA was measured by quantitative real-time polymerase chain reaction and Western Blot and standardised, automated immunohistochemistry were used to assess AR protein levels. The mean age at time of surgery was 61.8 and 30.9 months in boys without and with hypospadias, respectively. There was penile, coronal and sine hypospadias in seven (35%), nine (45%), and four (20%) boys, respectively. AR mRNA was significantly elevated in the prepuces of boys with hypospadias compared with boys without hypospadias, at a mean (sd) of 28.33 (5.39) vs 15.31 (1.85) (P = 0.013). Furthermore, the amount of AR protein was higher in boys with, compared with boys without hypospadias, at a mean (sd) of 133.25 (6.17) vs 100 (4.45) (P = 0.014). Different AR mRNA expression and protein levels seem to be an indication of an AR signalling defect as a cause in boys with hypospadias. Decreased AR DNA binding and functional capability may result in a compensatory up-regulation of both AR mRNA and protein. Further studies are necessary to perform structural analysis of the AR and to corroborate these preliminary findings.