Effects Of 2-methoxyestradiol On Growth, Invasion And Migration Of Mouse Melanoma B16 Cells

Objective: Malignant melanoma is a relatively common malignancy, occured in the skin and has a significantly higher capacity of invasion and migration. The long-term survival rate of metastatic melanoma is merely 5% and the incidence is increasing year by year. Nowadays, the key therapy of malignant melanoma is decreasing the risk behavior-solarization and early diagnosis and early treatments. Currently, the primary mean of treating malignant melanoma in situ is surgical excision. The effects of other anti-tumor therapies(immunotherapy,chemotherapy and radiation)are worse. The gene target therapy has a bright therapeutic perspective without extensive researches. To find a new effective treatment is extremely important. In recent years, many academics find that 2-methoxyestradiol has antitumor effects and induces apoptosis for many types of tumors, which plays an important role in inhibiting tumors occurrence and developments such as breast cancer,prostate cancer,gastric carcinoma and liver cancer. Now, 2-methoxyestradiol has been used as a clinical antitumor agent for breast cancer. The purpose of these trials is to observe the effects of 2-methoxyestradiol on proliferation, invasion, migration of mouse melanoma B16 cells,that can provide a new theoretical basis for the possibility of treatment by 2-methoxyestradiol on malignant melanoma.Methods:1 Cell cultureB16 cells were cultured in RPMI-1640 medium contained 10% fetal calf serum(including penicillin 100U/ml, streptomycin-containing 100μg /ml, PH7.3),which were placed in saturated humidity, 37 ℃, 5% CO2 Training Box.2 Observing morphological changes of B16 cells by inverted Microscope after 48 h.3 The MTT analysis method was to testify the proliferation of B16 cells treated with 2-methoxyestradiol in different concentrations. The cells untreated were regarded as the control group.4 The plate colony assay was used to testify the colony-formation ability of B16 cells treated with 2-methoxyestradiol in different concentrations for 2 weeks(10μmol/L,20μmol/L, 40μmol/L). The cells untreated were regarded as the control group.5 The Transwell assay was used to detect the capacity of invasion and migration of B16 cells treated with 2-methoxyestradiol in different concentrations. The cells untreated were regarded as the control group.6 The Wound healing assay was used to testify the capacity of migration of B16 cells treated with 2-methoxyestradiol at the concentrations of 10μmol/L for 24 h.The cells untreated were regarded as the control group.7 To analysis statistic dates use statistical software SPSS13.0Results:1 The morphological changes of B16 cells: Cells in the negative control group were spindle of growth well, transparent with good refractivity; cells treated with 2-methoxyestradiol became round. Floating cells and karyopyknosis were observed, along with the concentration increasing, the cells had became more and more.2 MTT assay showed that 2-ME of 10,20,40 μmol/L significantly inhibited the proliferation of B16 cells(F=920.78,P<0.05),with significant differences in the inhibitory effects among various durations of treatment with different concentrations of 2-ME( F = 6245.73,P< 0.05). There were significant interaction effects between drug concentration and treatment(95.0±11.4),(67.3±9.7)respectively. The differences between the various groups were significantly(P <0.05).4 The Transwell assay proved that the capacity of invasion and migration of B16 cells treated with 2-methoxyestradiol was inhibited compared with the control group(F=115.4 P<0.05, F=67.227 P<0.05).The cell numbers of invasion and migration between different groups were significantly difference(P<0.05).5 The Wound healing assay detected that the capacity of migration of B16 cells treated with 2-methoxyestradiol was inhibited compared with the control group.The cells in the control group almost migrated to all the scratches. Only some scratches area were occupied by cells in the10μmol/L experimental group.Conclusion:1 2-Methoxyestradiol could inhibit the proliferation of mouse melanoma B16 cell in a time-dose-dependent manner.2 2-Methoxyestradiol could inhibit the cloning ability of mouse melanoma B16 cell.3 2-Methoxyestradiol could inhibit the capacity of invasion and migration of mouse melanoma B16 cell.