## 5 Discussion  
### What Can Be the Targets for Treating Hair Graying?  
Premature or senile hair graying occurs when hair growth is relatively normal, but pigmentation is insufficient. Therefore, the most direct cause of hair graying is the decline in the viability and function of melanocytes that synthesize and supply melanin [164].  
MITF performs a central regulatory function in the differentiation and proliferation of melanocytes and melanin synthesis [69, 165]. Therefore, various factors can influence hair graying if they are upstream regulators of MITF, downstream targets of MITF, or interacting partners of MITF. The SCF/c-KIT, ET1/ETRB, WNT/Frizzled, and \(\alpha\)-MSH/MC1R signaling pathways are involved in MITF regulation [80]. Mutations and expression levels of MITF and several proteins that interact with it, such as SOX10 and IRF4, can affect the differentiation of MSCs and the death and function of melanocytes, leading to hair graying [70, 71]. Topical \(\alpha\)-MSH mimetic peptides have been proven effective in alleviating hair graying in patients [30, 31]. Topical placental sphingolipid increased MITF level in follicular melanocytes and black hair growth in age-onset gray-haired C57 BL/6] mice [43].  
Oxidative stress is caused by the overproduction of ROS or other pro-oxidants and a lack of antioxidants or antioxidant enzymes, resulting in oxidative damage to the key cellular molecules, such as nucleic acids, lipids, and proteins [89, 95]. Oxidative stress is both a cause and a consequence of aging and various diseases. Poor nutrition, lack of exercise, smoking, alcohol consumption, radiation, and air pollution can all increase oxidative stress, accelerating hair graying [85, 86, 87, 88, 89]. Oxidative stress due to X-ray irradiation, PUVA phototherapy, H\({}_{2}\)O\({}_{2}\), or hydroquinone can induce hair graying, which symptoms can be alleviated by various antioxidant strategies: inhibition of ROS generation, scavenging ROS using antioxidants or antioxidant enzymes, and increasing the antioxidant capacity of cells [96, 101, 156, 158].  
Stress hormones are the main triggers of hair graying [104, 105], while male hormones are one of the main triggers of alopecia [166]. An injection of resiniferatoxin causes pain stress and hair graying in animal models by stimulating norepinephrine secretion from the affected area [104, 105]. The norepinephrine-\(\beta\)2 AR-PKA signaling pathway mediates the pain stress responses in the body, including rapid differentiation and depletion of MSCs in the hair follicles, leading to hair graying [104, 105]. A natural alkaloid compound, rhynchophylline, was identified as a novel \(\beta\)2 AR blocker and its biological activity antagonizing the action of norepinephrine was verified in cells [159]. In animal models, rhynchophylline alleviated resiniferatoxin-induced hair graying [159]. Repeated hair plucking also caused hair graying in animal models probably via pain induction, and several EA-derived flavonoid compounds reduced the occurrence of white or gray hair [104, 153].  
Taken together, pharmacological approaches enhancing the expression and activity of MITF in melanocytes or effectively mitigating the negative effects of oxidative stress and stress hormones are expected to provide useful strategies for treating hair graying.