Shikonin Inhibits The Proliferation Of Middle Ear Cholesteatoma Epithelial Cells By Down-regulating The Expression Of AnxA2

Objective: To study the effect of shikonin on the proliferation and apoptosis of human immortalized keratinocyte cell line(HaCat cells)and the effect on the expression of Annexin A2(AnxA2),and to explore the inhibitory effect of shikonin on the proliferation of middle ear cholesteatoma epithelial cells and related molecular mechanisms,and lay the experimental and theoretical foundation for the related mechanism of shikonin in the treatment of middle ear cholesteatoma.Methods: Shikonin at different concentrations(0,1,5,10umol/m L)was used to interfere with HaCat cells.Observe the changes of cell morphology under a microscope;observe the migration ability of cells in different intervention groups by scratch experiment;detect cell cycle and apoptosis by flow cytometry;CCK-8 method was used to detect the effects of different concentrations of shikonin on HaCat cell proliferation inhibition and apoptosis induction;RT-PCR was used to detect the m RNA expression of AnxA2 in different intervention groups;Western-blot detects the expression of AnxA2 protein in different intervention groups.Results: Shikonin can make HaCat cells lose their original cell morphology,make the cells shrink,round,and decrease adherence.As the concentration of shikonin increases,cell changes become more obvious;Shikonin can block HaCat cell cycle,with the increase of shikonin concentration,the higher the proportion of HaCat cells in G1 phase and the lower the proportion in S phase,the difference is statistically significant(P<0.05);shikonin can Inhibit the migration ability of HaCat cells,and as the concentration of shikonin increases,the migration ability of HaCat cells becomes worse;shikonin can induce apoptosis of HaCat cells,and as the concentration of shikonin increases,the The apoptotic rate gradually increased,and the difference was statistically significant(P<0.05).Shikonin at different concentrations can inhibit the proliferation of HaCat cells in a dose-dependent manner,and the difference is statistically significant(P<0.05),and shikonin at a concentration of 10umol/m L has a significant inhibitory effect on the proliferation of HaCat cells(P<0.01).RT-PCR method detected the m RNA expression of AnxA2 in HaCat cells of different intervention groups.The results showed that with the increase of shikonin concentration,the m RNA expression level of AnxA2 decreased in a dose-dependent manner,and the difference was statistically significant(P<0.05).The Western-blot method showed that as the concentration of shikonin increased,the expression level of AnxA2 protein gradually decreased.Conclusion: Shikonin can inhibit the proliferation of HaCat cells in a dose-dependent manner,and its mechanism may be related to the down-regulation of AnxA2 expression.