Iris de Nie

86 C H A P T E R 6 MATERIALS AND METHODS Study population and clinical data collection For this study, we used orchiectomy specimens of transgender women who underwent bilateral orchiectomy combined with vaginoplasty at the Center of Expertise on Gender Dysphoria of Amsterdam UMC between 2006 and 2019. All participants provided written permission for the use of their body material and clinical data for research purposes. The Ethical Review Board of the Amsterdam UMC, location VUMC provided approval for conducting this study (METC2014322). A total of 788 transgender women were identified. Data on medical history, age and Tanner stage at start of medical treatment, documented hormone use, date of gGAS, alcohol consumption, smoking, drug use, BMI at time of gGAS, and last known serum hormone levels before gGAS, were collected from the medical files. Transgender women were categorized according to age and Tanner stage at initiation of medical treatment (Tanner stage 2-3, Tanner stage 4-5, or ≥ 18 years). Transgender women operated before 2017 discontinued GAHT four weeks prior to surgery, because of a presumed increased risk of perioperative thrombosis. As evidence suggested this risk is negligible, GAHT is continued in the perioperative period since July 2017. Six subgroups were created based on Tanner stage/age at start of medical treatment and continuation/discontinuation of GAHT prior to gGAS. People with an unknown age or Tanner stage at time of initiation of medical treatment were excluded. Other exclusion criteria were hard drug use, cryptorchidism, a medical history of receiving chemotherapy, or genetic disorders which can all possibly impair spermatogenesis. Lastly, since the vast majority used estrogens combined with either triptorelin, or cyproterone acetate, people who used estrogen monotherapy and those who used spironolactone as anti-androgenic treatment were excluded to create a homogeneous study population. A maximum number of 80 transgender women were enrolled per group as this was deemed sufficient to answer the study questions. A random sample was drawn from groups that exceeded 80 individuals using STATA Statistical Software, version 15.1 (Statacorp, College Station, TX, USA). In total, 263 transgender women were selected for inclusion in the study cohort. Testicular tissue preparation and analysis Preparation for histology Testicular tissue was obtained from the biobank of the Pathology department of Amsterdam UMC, where orchiectomy specimens, obtained during gGAS, were stored after histopathological analysis for clinical purposes. Upon arrival at the Pathology department, the orchiectomy specimens were fixed in 4% w/v paraformaldehyde and embedded in paraffin. For this study, seven slices of 5 μ m thickness of one testicle were sectioned and mounted on microscope slides. From one slide of each specimen paraffin sections were deparaffinized and subsequently stained with Mayer’s haematoxylin and eosin, and at least one other slide was used for immunohistochemistry to confirm germ cell subtypes.