ESPE Abstracts

Introduction: Hypospadias is often associated with reduced testosterone synthesis or action. High rates of complications are reported with surgical repair of hypospadias, including wound dehiscence. Boys with hypospadias have previously been demonstrated to have increased reactive oxygen species (ROS) compared to healthy controls and there is a known link between male hypogonadism and oxidative stress. It is not clear what effect ROS has on wound healing in boys with hypospadias.

Aims: To identify if cell migration and proliferation in genital skin are altered in boys with hypospadias, and whether this can be altered by antioxidants.

Methods: Genital skin (GS) samples were collected from boys undergoing hypospadias repair (cases) or routine circumcision (controls) for GS fibroblast culture. Cells were seeded at a density of 100,000 cells/well using a haemocytometer and grown until 80% confluence. A sterile pipette tip was used to scratch a wound. Cells were imaged using an EVOS XL Core microscope immediately after the wound was made and 48 hours later, in the presence/absence of ROS scavengers, N-acetylcysteine (NAC) or Tempol. Cell migration was determined using ImageJ software. Cell proliferation was measured using a commercial Cell Count Kit-8 (Abcam, UK).

Results: Twenty four cases (median age (range) 1.8 (1.2, 6.3) years) and 28 controls (median age 1.6 (1.2, 6.1) years) were recruited. Of the boys with hypospadias, 14 (58%) had distal hypospadias, 9 (38%) had proximal hypospadias and 1 (4%) had midshaft hypospadias. The median External Masculinisation Score of cases was 10 (3, 11). There were no statistically significant differences in endocrine biochemistry between the 2 groups at the time of surgery. Genetic testing had been performed in all proximal hypospadias cases, with no variants identified. Boys with hypospadias had impaired cell migration with reduced % wound closure at 48 hours (2.0 fold, P<0.0001) and reduced cell proliferation (1.3 fold, P=0.01). External Masculinisation Score was positively correlated with % wound closure (r= 0.5, P<0.0001) and cell proliferation (r= 0.3, P=0.002). There were no statistically significant correlations between birthweight, gestation at birth or endocrine biochemistry. Exposure to both NAC and Tempol improved wound closure (1.9 fold, P=0.01, and 1.5 fold, P=0.02 respectively) and cell proliferation (1.5 fold, P=0.02 and 1.4 fold, P=0.05 respectively).

Conclusions: There is an association between wound healing and virilisation of the external genitalia in boys. ROS scavengers improve cell migration and proliferation in boys with hypospadias and could represent a therapeutic solution for reducing surgical complications.