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53 A COHORT STUDY ON FACTORS IMPAIRING SEMEN QUALITY IN TRANSGENDER WOMEN 4 INTRODUCTION Many young people with gender dysphoria – distress caused by an incongruence between the sex assigned at birth and the gender identity – seek medical treatment to align their physical characteristics with the identified gender. 70 This medical treatment involves gender- affirming hormonal treatment (GAHT), often followed by gender-affirming surgery (GAS). 93 For transgender women – people assigned male at birth with a female gender identity – GAHT generally consists of a combination of antiandrogens and estrogens. 2 After at least one year of GAHT, transgender women may choose to undergo GAS which may involve bilateral orchiectomy often combined with vaginoplasty. 93 Although this treatment significantly improves quality of life, the loss of reproductive function is an often unwanted consequence. 60,73,94 The use of gender-affirming hormones severely impairs semen quality which might not, or only partially, recover after cessation of this medication. 39,95 After bilateral orchiectomy, reproductive loss is permanent. Therefore, it is important that (future) desire for children and the options for fertility preservation are discussed and offered prior to the start of gender-affirming treatment. Cryopreservation of ejaculated spermatozoa is preferred, as it is physically noninvasive and provides the best semen quality. However, our previous retrospective study showed that even before initiating GAHT, semen quality in transgender women is decreased compared to cisgender men – birth assigned males who also identify as male – and in a substantial percentage of semen samples, quality is lower than WHO reference values for human semen. 10,11,96 Therefore, it is worth identifying lifestyle interventions which may improve the pre-freeze quality. Previously we were not able to provide an explanation for the impaired semen quality in transgender women. Factors known to have a negative influence in the general population like smoking, alcohol abuse or obesity, were not responsible for the overall decreased semen quality in our cohort. 96 However, semen quality might also be affected by habitual behavior more typically observed in transgender women, such as the desire to hide their testicles. Scrotal hyperthermia, as a result of wearing tight undergarment or bringing the testicles into the inguinal canal (tucking), may negatively influence sperm motility and concentration. 88-92 Furthermore, genital dysphoria may result in a low frequency of ejaculation which may negatively impact sperm motility. Although these behavioral factors are often assumed to be explanatory for the impaired semen quality, their frequency and effect on semen quality in transgender women have never been studied. The purpose of the current study is to obtain data on all factors with a potential negative impact on semen quality in transgender women and further validate the findings from our previous study.