Naturally occurring Ngn2 promoter activators from Butea superba

Neurogenin2 (Ngn2), an activator-type bHLH transcriptional factor, promotes differentiation of neural stem cells into neurons by transcription of pro-neural genes. To find neural stem cell accelerators from the extract library of natural resources, we used a two-step screening including a Ngn2 promoter reporter gene screening and differentiation assay screening of neural stem cells. A reporter gene assay that can detect Ngn2 promoter activity by luciferase expression was constructed using C3H10T1/2 cells. Using this primary cell-based screening, Butea superba was found to include Ngn2 promoter activators from our tropical plant extract libraries. Bioassay-guided fractionation of this plant extract led to the isolation of 18 Natural Products, including pterocarpans and isoflavonoids. Dehydromaackiain (1), formononetin (6), ()-variabilin (13), ()-medicarpin (14), rothindin (17) and ononin (18) showed 1.8–2.8 times higher Ngn2 promoter activity at 5 mM compared with control. Of active natural compounds, 30-methoxydaidzein (3) showed promotion of neurite outgrowth of C17.2 in a secondary screen. 30-Methoxydaidzein (3) increased mRNA expression of pro-neural transcriptional factors (Ngn2, Ngn1, NeuroD2), a mature neuron-specific Enzyme GAD1 and a pro-neural neurotrophic growth factor neurotrophin 3 (NT3) in C17.2 neural stem cells.