ABSTRACT
Objective To evaluate the effect of latanoprost on cell proliferation of and melanogenesis in human epidermal melanocytes,and to explore its mechanism.Methods Latanoprost was added into the 254 medium to prepare latanoprost solutions at different concentrations of 10-5,10-6 and 10-7 mol/L.In vitro cultured human epidermal melanocytes were divided into 4 groups to be cultured with media containing no latanoprost (control group) or 10-5,10-6 and 10-7 mol/L latanoprost for 48 hours.Cell counting kit-8 (CCK8) assay was performed to evaluate the proliferative activity of melanocytes,dopa oxidation assay to estimate the activity of tyrosinase.Sodium hydroxide (NaOH)-lysis method was used to determine the content of melanin,and Masson-Fontana staining to observe the number and distribution of melanin granules.Westernblot analysis and real-time fluorescence-based quantitative PCR were performed to determine the protein and mRNA expression of melanogenesis-related genes including microphthalmia-associated transcription factor (MITF),tyrosinase (TYR) and tyrosinase-related protein 1 (TYRP1).Comparison among the 4 groups and multiple comparisons were done by using one-way analysis of variance and least significant difference (LSD)-t test.Results Compared with the control group,the 10-6-,10-5-mol/L latanoprost groups showed significantly increased proliferative activity of melanocytes (1.064 ± 0.172 and 1.078 ± 0.080 vs.0.784 ± 0.015;t =3.289,3.454 respectively,both P ＜ 0.05),increased activity of tyrosinase (0.510 ± 0.017 and 0.454 ± 0.009 vs.0.355 ± 0.041;t =6.139,3.939 respectively,P ＜ 0.01 or 0.05),and increased content of melanin (t =7.232,5.967,both P ＜ 0.01).However,there were no significant differences in the proliferative activity of melanocytes,activity of tyrosinase or content of melanin between the 10-7-mol/L latanoprost group and control group (all P ＞ 0.05).Masson-Fontana staining showed more and darker melanin granules on melanocyte dendrites in the 10-5-,10-6-,10-7-mol/L latanoprost groups than in the control group,and the color of melanin granules changed from light brown to black brown along with the increase in the concentration of latanoprost.The mRNA expression of MITF increased along with the increase in the concentration of latanoprost (P ＜ 0.01),and the protein expression of MITF wassignificantly higher in the 10-6,10-5-mol/L latanoprost groups than in the control group and 10-7-mol/L latanoprost group (all P ＜ 0.01).The 10-6-mol/L latanoprost group showed significantly increased mRNA and protein expression of TYR and TYRP1 compared with the control group,10-7-,10-5-mol/L latanoprost groups (all P ＜ 0.01).Conclusion Latanoprost can increase the proliferation of human epidermal melanocytes,and promote tyrosinase activity and melanogenesis likely by enhancing the mRNA and protein expression of MITF,TYR,TYRP1.