Endocrine Abstracts

Introduction: Mitochondrial uncoupling proteins (UCPs) are transmembrane channels present in the inner mitochondrial membrane whose function is the transport of protons and small substrates between the intermembrane space and the mitochondrial matrix. UCPs are key regulators of cellular metabolism, including mitochondrial function, oxidative phosphorylation, and reactive oxygen species (ROS) production. UCPs’ dysfunction has been associated with the onset of metabolic diseases, such as obesity and diabetes mellitus. In men, metabolic diseases are closely linked to hypogonadism and lower testosterone production in Leydig cells, however, the molecular mechanisms are still poorly understood. The dysfunction of UCPs could play a role in the crosstalk between metabolic diseases and hypogonadism but their expression and function in Leydig cells is still unknown.

Aim of the study: The aim of this study was to identify the expression of UCP1, UCP2, and UCP3 in mouse Leydig Cells (mLCs). Additionally, the influence of UCPs on the mitochondrial function was assessed, through its inhibition by genipin, a selective UCP inhibitor.

Materials and methods: Culture of mouse Leydig cells (BLTK1 cell line) were established (n=10). Total RNA was extracted and Ucp1, Ucp2, and Ucp3 mRNA expression were assessed by RT-PCR. UCP1-3 protein expression was determined by Western Blot and immunofluorescence. To assess UCPs function, mLCs were treated with genipin (0.5, 5, 50 and 100 µM). After 24 h treatment, cellular proliferation and viability were evaluated. Mitochondrial function was accessed by Seahorse XF Cell Mito Stress assay kit.

Results: We were able to identify both the mRNA and protein expression of UCP1-3 in mLCs. The inhibition of UCPs decreased cellular proliferation and viability of mLCs. Additionally, the mitochondrial function was severely affected by UCPs’ inhibition.

Conclusion: In this work, we were able to identify for the first time, to the best of our knowledge, the expression of UCP1-3 in mLCs. Our data highlight that UCPs are modulators of mLCs mitochondrial function, suggesting a potential regulatory role of UCPs on steroidogenesis. Taken together, the discovery of UCPs in Leydig cells opens the path for future studies focusing on the molecular mechanisms responsible for the metabolic diseases-induced male hypogonadism.