Chlorocholine chloride disrupts differentiation of rat stem/progenitor Leydig cells: A combined in vivo and in vitro study by Leydig cell regeneration model.

Chlorocholine chloride (CCC), a widely-used plant growth regulator, impairs testosterone biosynthesis, but the mechanism remains elusive. The Ethane dimethane sulfonate (EDS)-induced rat Leydig cell regeneration model was adopted to investigate the effect of CCC on Leydig cell development and testosterone levels. Rats received 75, 137.5, or 200 mg/kg bw CCC from post EDS d7 to d28 or d56. At d28, CCC exposure inhibited stem/progenitor Leydig cell differentiation. At d56, CCC exposure affected the maturation status of adult Leydig cells, reduced expression of StAR, HSD3B1, and serum and testicular testosterone levels. Disrupted hypothalamic-pituitary-testis axis and activated PI3K/AKT pathway might participate in the CCC-induced Leydig cell regeneration retardation. In vitro, 50 μg/mL CCC inhibited Leydig cell differentiation and testosterone levels. Collectively, CCC delays Leydig cell development in vivo and in vitro by suppressing differentiation, leading to lower mature Leydig cells and reduced testosterone levels.