Study On The Effect Of TSPO Ligand FGIN-1-27 On Melanin Synth Esis And Its Mechanism

The melanin has great significance to the human skin,it can act as a protective barrier against external stimuli and maintain the normal life activities of body.However,the abnormal accumulation of melanin can cause skin problems,and even lead to skin cancer.In the current,most whitening products are added with whitening active ingredients,which play a role in inhibit melanogenesis and contribute to melanin metabolism.Although a large products of whitening on the market,most of them have problems such as cell injury or poor effect.Therefore,it is necessary to explore more effective and safe whitening active ingredients.The translocator protein(TSPO)is an 18 k D mitochondrial membrane protein,it has multiple physiological functions,including cell growth and proliferation,specialized steroidogenic,cellular immunity and mitochondrial respiratory apoptosis.Prior studies have noted that PK11195 has an inhibitory effect on the melanin synthesis,indicating that TSPO can be used as a candidate target for the treatment of pigmented skin diseases,and the small-molecule ligands bind to it may have the ability to regulate pigment metabolism.FGIN-1-27 as a ligand of TSPO,has demonstrated pro-apoptotic,anti-anxiety and steroidogenic activities,it remains uncertain that regulate melanin synthesis.Our research aims to study the effect of FGIN-1-27 on tyrosinase activity in vivo and in vitro,and the effect on different melanocytes and UVB-induced pigmentation of guinea pig skin,to verify whether it has the function of regulating melanin synthesis and investigates the underlying mechanism.Synthesis of FGIN-1-27: Beginning with 3-(4-Fluorobenzoyl)propionic acid as the starting material,FGIN-1-27 was synthesized,and its structure was characterized by NMR.FGIN-1-27 and TSPO protein docking: use the CDOCKER module in Discovery Studio software for docking processing,select 8 different TSPO protein ligands,through a range of ligands and protein processing,ligand-receptor binding,Check the energy value.The results show that the binding stability of FGIN-1-27 and TSPO protein is the highest.Direct effect of FGIN-1-27 on tyrosinase activity: the detection model of tyrosinase inhibitor is established,and the optimal conditions are selected: the concentration of L-tyrosine is 2.5 mmol/L,mushroom tyrosine The concentration of the enzyme is 2000 U/m L,and the reaction time is 20 min.This model is used to determine the activity of mushroom tyrosinase to detect whether FGIN-1-27 has inhibitory activity and compare it with other whitening products.The results show that FGIN-1-27 has no direct inhibitory effect on mushroom tyrosinase activity,and FGIN-1-27 is not a direct inhibitor of tyrosinase.Effect and mechanism of FGIN-1-27 on melanin synthesis: FGIN-1-27 has no effect on cell viability and has no cytotoxicity by MTT cell viability detection.The effect of FGIN-1-27 on melanogenesis was detected by Na OH lysis method and Masson-Fontana ammonia silver staining method.The results showed that FGIN-1-27 can inhibit melanocyte stimulating hormone(a-MSH),endothelin-1(ET-1)and OAG(DAG analog)induced melanin synthesis.The effect of FGIN-1-27 on cell melanin synthesis and related regulatory proteins was determined by Western blot and immunofluorescence quantitative experiments.The results indicate that FGIN-1-27 mainly acts by inhibiting the PKA,PKC and MAPK pathways.Effect of FGIN-1-27 on guinea pig pigmentation model: A UVB-induced guinea pig pigmentation model was established,and the control method of administration was adopted.Through direct observation,the change of guinea pig skin color value was measured,Masson-Fontana ammonia silver staining method and immunohistochemical staining verified the effect of FGIN-1-27 on the local skin melanin production and the number of melanocytes in guinea pigs.The results showed that FGIN-1-27 reduced UVB-induced skin pigmentation in guinea pigs,but did not reduce the number of melanocytes.