Hedgehog signalling in solitary motile cilia: detection and role of hedgehog factors in sperm functions and in vitro fertilization

The Hedgehog (Hh) signalling pathway controls cellular functions important to organ development and homeostasis. While previously thought to be restricted to non-motile primary cilia, a study indicated that Hh modulates cAMP concentration and beating frequency of airway motile cilia. Since systemic blockade of Hh signalling reduces sperm motility in vivo, we hypothesised that Hh agonists present in the environment surrounding the sperm could directly modulate sperm functions. The presence and localisation of Hh factors, including PATCHED1 receptor, GLI3, and SMOOTHENED, were investigated in spermatozoa from murine, bovine, and human species. Following sperm incubation under capacitating conditions with either Hh agonist (smoothened agonist; SAG) or inhibitor (Cyclopamine), functional sperm parameters were measured, including motility, acrosome integrity, cell viability, intracellular calcium levels, cAMP concentrations, and in vitro fertilisation outcome. Our study demonstrates that Hh signalling pathway regulates sperm motility and acrosome reaction, contributing to fertilisation success. This report proposes new comprehensive signalling mechanisms that control male reproductive functions.