The testicular organoid as a model to study the impact of endocrine-disrupting chemicals on steroidogenesis.

A decline in sperm count worldwide has been observed over the last decades. Exposure to xenobiotics such as environmental pollutants, drugs, food additives and synthetic polymers exhibiting endocrine-disrupting activities is considered a significant contributing factor. Addressing this decline necessitates the development of an effective human testicular model to reliably identify male fertility disruptors. In this context, we investigated the potential of human testicular organoids (TOs) as a promising in vitro approach to mimic steroidogenesis. TOs were derived from testicular tissue obtained from six adult transgender patients, and cultured up to 63 days. To characterize steroidogenesis in TOs, transcriptome profiling with microarray, qRT-PCR and immunostaining were performed to assess the expression of key markers for Leydig cells and the steroidogenic pathway. Testosterone and estrogen production were measured in the culture medium of TOs by means of an electrochemiluminescence assay. The presence of Leydig cells and the expression of steroidogenesis-specific components in transgender TOs (n=3) were confirmed at both the gene and protein levels. The secretory function of Leydig cells tended to peak after two weeks, with an average testosterone concentration of 646.8620.1 ng/mL at day 14 (n= 6), followed by a gradual reduction and stabilization, reaching a concentration of 128.2±74.2 ng/mL after one month (n=6). Testosterone production persisted for up to 63 days, with a concentration of 73.3± 51.9 ng/mL (n=6). Also, estrogen was measured at day 14 and reached a concentration of 5.4± 2,2 ng/mL (n= 5). TOs harbor functional Leydig cells, expressing all major factors important for steroidogenesis and producing hormones such as testosterone and estrogen. This makes TOs a promising model for assessing the impact of endocrine-disrupting chemicals (EDCs) on steroidogenesis, facilitating the examination of EDC effects on male fertility. TOs hold promise for translating research findings into clinical implications or regulatory actions as a New Approach Methodology, contributing to the advancement of reproductive toxicology knowledge and guiding preventive measures.