Stigmasterol promotes neuronal migration via reelin signaling in neurosphere migration assays

Stigmasterol (ST) is a multifunctional phytosterol and is found in diverse food. In our previous transcriptomics study, we found ST upregulated migration-related genes. In the present study, we carried out in vitro neurosphere migration assays to investigate the effects of ST on neuronal migration. For this purpose, neurospheres were produced by culturing rat (Sprague-Dawley) E14 cortical neurons. The addition of ST (75 μM) to culture medium increased not only the numbers of migratory neurons by 15% but the distance of movement up to 120 μm from the centers of neurospheres as compared to vehicle cultures. Immunocytochemistry and immunoblotting showed ST upregulated the expressions of Reelin (Reln) and its downstream signaling molecules like phospho-JNK (c-Jun N-terminal kinase), doublecortin (DCX) and dynein heavy chain (DHC) in migratory neurons. Furthermore, in silico molecular docking simulation indicated that ST interacts with Relin receptor ApoER2 by forming a hydrogen bond with Lys2467 and other van der Waals interactions. Taken together, our study shows that ST upregulates Reln signaling and promotes neuronal migration and suggests that ST supplementation is considered as a potential means of treating migration-related CNS disorders.