Impact of a supernumerary X-chromosome on testicular somatic cell function

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Klinefelter syndrome (KS; 47,XXY) is the most frequent sex chromosomal aberration in men causing hypogonadism and infertility and is routinely treated with androgens. Up to 50% of patients show remnant spermatogenesis, which might be destroyed by testosterone (T) administration.The somatic testicular cells are also affected: Leydig cell (LC) hyperplasia is present and although intratesticular T levels are normal, serum T levels are insufficient. Impaired testicular vascularization is likely contributing to the endocrine disbalance. XXY Sertoli cells (SC) offer a niche and support for differentiating germ cells (GC), resulting in focal spermatogenesis but their physiology is affected by the disorder. Peritubular myoid cells (PMCs) are also functionally important but their role in the testicular degeneration in KS is likely underestimated. Our overarching aim is to delineate the testicular somatic cell function in the presence of a supernumerary X chromosome. KS affects the patients systemically and we will examine such effects using our 41 XXY* mouse model. We collected clinical data, DNA and testicular biopsies from several hundred patients and controls, enabling translation of results from the model. Even elevated Luteinising Hormone (LH) levels are stimulatory insufficient, likely due to hampered testicular transport of the large LH molecule. We hypothesize that LH receptor (LHR) stimulation by an agonist could improve LH action, enhance steroid response and normalize serum T. Previously, isolated LCs from XXY* mice and controls were treated: We found that i) the agonist is highly active and ii) XXY* LCs responded stronger. We confirmed this in murine 41,XXY* testicular organ culture indicating that the agonist can restore the endocrine signaling, a hypothesis to be challenged in vitro (human testicular tissues) and in vivo (41,XXY* mice). The expected results can enable a new treatment stimulating endogenous T production and protecting focal spermatogenesis. Single cell RNA sequencing (scRNAseq) indicated SCs to be involved in testis degeneration in KS. PMCs play a role in the maintenance of focal spermatogenesis and accelerated fibrosis. We hypothesize that PMCs and SCs are affected by the condition, resulting in a fading “window of opportunity” to foster remnant spermatogenesis. We will challenge this by intratesticular transplantation of donor GCs from eGFP mice, mimicking focal spermatogenesis, scRNAseq transcriptomic approaches, and subsequent bioinformatics assessment. The detected candidates will be translated and validated at the protein level in testis biopsies from patients with focal spermatogenesis. We will analyze PMCs and SCs during different phases of reproductive life and with regard to present or absent somatic – GC interactions. We will gain insights on chromosomal aberration effects on testicular somatic cell function and clinically highly relevant data for the development of improved therapies for KS.