The Study Of The Effect Of Nutlin-3on Antitumor And Antiinvasion In Human Melanoma A375Cells

Objectives:1.To study the effect of nutlin-3in human melanoma A375cells proliferation, arrest cell cycle and apoptosis in vitro and to establish its foundation in aspect of antitumor activity in melanoma.2. To evaluate the effect of nutlin-3in human melanoma A375cells in invasion, and to study its possible mechanism of antiangiogenesis.Methods:1. MTT assay was performed to study the effect of nutlin-3in human melanoma A375cells proliferation.2. Inverted lapse microscope was utilized to observe morphological changes of human melanoma A375cells treated with nutlin-3.3. Flow cytometer was used to test the effect of nutlin-3in human melanoma A375cells cycle arrest.4. Flow cytometer was used to test the effect of nutlin-3in human melanoma A375cells apoptosis.5. To detect the antiinvasion ability of nutlin-3in human melanoma A375cells by transwell.6. Western blot was performed to study the effect of nutlin-3inducing the expression of P53and VEGF-A in human melanoma A375cells.Results:1. The results of MTT assays demonstrated that different dose (1.25μM,2.5μM,5μM,10μM) of nutlin-3suppressed the human melanoma A375cells proliferation.They had the dose-dependent relationship.2. Under inverted lapse microscope, human melanoma A375cells were spindle-shaped adherent growth, cell debris drifted in the liquid level, the number continued to decline, after treated with different concentrations (2.5μM,5μM,10μM) of nutlin-3.3. The results of flow cytometer assay showed that different concentrations (2.5μM,5μM,10μM) of nutlin-3could significantly induce the cell cycle arrest of the human melanoma A375cells in the dose and time-dependent manner.4. The results of flow cytometer assay showed that different concentrations (2.5μM,5μM,10μM) of nutlin-3could significantly induce the apoptosis of the human melanoma A375cells.5. The transwell and variation of optical density indicated that different dose (2.5μM,5μM,10μM) of nutlin-3could obviously inhibit human melanoma A375cells attaching to ECM and had a positive correlation between the invasion inhibition ratio and the dose of nutlin-3.6. The results of western blot showed that different concentrations (2.5μM,5μM,10μM) of nutlin-3increase the expression of P53in human melanoma A375cells.7. The results of western blot showed that different concentrations (2.5μM,5μM,10μM) of nutlin-3decrease the expression of VEGF-A in human melanoma A375cells.Conclusion:1. Different dose (1.25μM,2.5μM,5μM,10μM) of nutlin-3can inhibit the proliferation of human melanoma A375cells, arrest cell cycle and induce apoptosis, it has an obvious antitumor activity in the melanoma and it’s deserved to study and research extremely.2. Different dose (2.5μM,5μM,10μM) of nutlin-3can suppress the human melanoma A375cells invasion, and its mechanism is possible concerned with downregulation expression of MMP-2.3. Different dose (2.5μM,5μM,10μM) of nutlin-3can induce P53restoration, downregulate the expression of VEGF-A in human melanoma A375cells, it has very good activity in repressing angiogenesis, which is deserved to study and research extremely.