The Effects Of Endothelin3(EDN3) On Proliferation And Melanin Synthesis Of Alpaca Skin Melanocytes In Vitro

The hair color of mammalian is decided by the amount, the types and the distribution of melanins produced by melanocytes. Endothelin takes part in the cell growth factors strongly and promoting the cellular mitosis. EDN3plays an important role in the growth of melanocytes from neural crest cellular precursor. Although, the effects of EDN3on melanoblast, melanocytes and melanocytoma has been known, the mechanism of EDN3in melanocytes is not known well. Here, the proliferation of melanocyte, the production of melanin in melanocytes and the expressions of associated hair color genes were examined following the addidion of EDN3to the medium of alpaca melanocyte culture in vitro, to discover the mechanism of EDN3in the formation of hair color.In the experiment, added different dozes of EDN3(10-9mol/L、10-8mol/L、10-7mol/L) in the primary cultured4th alpaca skin melanocytes.After72h with the addition of EDN3, the contents and the results are as following:1. The control group of alpaca skin melanocytes were two or three dendritic and the number of dendrites were relatively lower. Compared with the control group, the amount of the cells and synapse of the melanocytes was increased and the cytoplasm of the melanocytes grew bigger in the experiment group, especially the group with EDN3of10-8mol/L.2. Examined the proliferation of the alpaca melanocytes by MTT method. Compared with the control group, EDN3of the dose of10"9mol/L stimulated the proliferation of the melanocytes and the dose of10-7mol/L has the opposite effect, but the dose of10-8mol/L has the best effect (P<0.05)3. The real-time PCR showed that the expression of EDNRB、KIT、MITF、TYR mRNA was3.21,3.58,3.08and2.63fold changes (P<0.05), respectively, in the experiment group with EDN3addition of10-8mol/L, compared with the control group.4. Western blotting results showed that the expression of EDNR、KIT、MITF and TYR was2.34,2.22,3.46and1.59fold changes (P<0.05) in melanocytes in experiment group with EDN3addition of10-8mol/L, compared with the control group. 5. Examined melanins synthesis in alpaca melanocytes by UV spectrophotometry. The. melanins production in alpaca melanocytes cultured in vitro was increased in melanocytes with EDN3addition of10-8mol/L, melanin synthesis was1.44fold changes (P<0.05), compared with the control group.Above all, EDN3promoted the proliferation of alpaca melanocytes by up-regulated the expression of transcription and protein in EDNRB、KIT、MITF and TRY in the cells, furthermore it promoted melanin synthesis in melanocytes.