Induction of insulin‐like growth factor‐I by cepharanthine from dermal papilla cells: A novel potential pathway for hair growth stimulation

Cepharanthine (CEP) is a biscoclaurine alkaloid extracted from Stephania cepharantha and has been commonly prescribed for internal use in Japan for more than 40 years to treat a variety of diseases including alopecia areata. Previously, it was reported that topical CEP stimulated hair growth of C3H/HeN mice and 0.1 μg/mL CEP stimulated the growth of mouse dermal papilla cells (DPC)1 in vitro, suggesting stimulation by CEP on hair growth through direct effects on DPC. Here, we study the molecular mechanism of CEP stimulation of human balding DPC. The DPC obtained at passages 4–6 from a male androgenetic alopecia (AGA) bald frontal scalp were cultured on a 12-well plate in Dulbecco's modified Eagle's medium (Nissui Pharmaceutical, Tokyo, Japan) supplemented with 10% charcoal-treated fetal calf serum (JRH Biosciences, Lenexa, KS, USA), penicillin (50 units/mL) and streptomycin (50 μg/mL) at 37°C in a humidified atmosphere of 95% O2 and 5% CO2. At subconfluency, 0.1 and 1.0 μg/mL CEP or ethanol mock was added to the culture and 24 h later the cells were harvested. RNA was extracted from the cells and subjected to semiquantitative reverse transcription polymerase chain reaction for well-known hair growth regulators such as insulin-like growth factor-I (IGF-I), fibroblast growth factor-2, leptin, interleukin (IL)-1α, IL-6, IL-8, transforming growth factor (TGF)-β1, vascular endothelial growth factor (VEGF)-A, hepatocyte growth factor (HGF) and tumor necrosis factor (TNF)-α as described previously.2 As a result, only IGF-I mRNA increased by 0.1 and 1.0 μg/mL CEP compared with the mock treatment (Fig. 1a) and the effect was enough at the concentration of 0.1 μg/mL. We could not detect mRNA of VEGF-A, HGF and TNF-α using this line of DPC. Then, to confirm this effect by CEP at protein level, we examined the effect of CEP on IGF-I secretion in the conditioned media using the three primary culture DPC lines from three male AGA patients. At subconfluency, in 5 mL medium on the 6-cm dishes, we added 0.1 μg/mL CEP to the culture and continued the incubation for 72 h. Then, the conditioned media were harvested and subjected to enzyme-linked immunoassay (ELISA) for IGF-I using an Human IGF-I Quantikine ELISA Kit (DG100; R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions. As a result, 0.1 μg/mL CEP significantly increased IGF-I secretion by 1.978 ± 0.566 fold compared with mock controls (Mann–Whitney U-test, P < 0.05) (Fig. 1b). From these findings, CEP stimulates hair growth through potentiating IGF-I production from DPC. In this study, we found that IGF-I is a potential target for CEP as a hair growth stimulator. IGF-I is an androgen-inducible positive growth mediator for beard growth stimulation3 and an increased expression of IGF-1 mRNA in the DPC is reportedly associated with patient response to finasteride,4 indicating that IGF-I can be a key factor in therapeutic responsibility of AGA. Therefore, our data here suggest that CEP is a hopeful reagent for AGA. In this study, the concentration of CEP was 0.1 and 1.0 μg/mL but it is extremely higher than serum concentration after taking 10 mg oral CEP (1 ng/mL),5 suggesting that topical application of CEP is more feasible for treating AGA or other hair loss diseases through the stimulatory effect on IGF-I production shown here. Neither of the authors has a financial or commercial interest.