Expression And Significance Of H3K27me3 In Middle Ear Cholesteatoma

Objective:To compare the expression of histone H3K27me3 in the epithelium of middle ear cholesteatoma and the corresponding epithelium of external auditory canal,and detect the factors that may be related to the abnormal expression of histone H3K27me3,so as to explore its significance in the formation and progress of middle ear cholesteatoma,and try to explain the pathogenesis of middle ear cholesteatoma in the epigenetic level.Methods:1.From October 2018 to June 2019,35 patients with cholesteatoma otitis media were collected from the Department of Otolaryngology,Affiliated Hospital of Qingdao University.The fresh cholesteatoma epithelium removed during the operation was selected as the experimental group,and the normal external auditory canal epithelium corresponding to the same patient was selected as the control group.Western blot was used to detect the expression of H3K27me3 in 35 cases of cholesteatoma.2.At the same time,25 cases in the experimental group and 25 cases in the control group of the same patient were collected,fixed by OCT embedding agent in tissue frozen section,and the expression of H3K27me3 was localized and semi quantitatively detected by immunofluorescence staining after frozen section.3.The expression of EZH2,EZH1,KDM6A,TGM1 and IVL gene mRNAin the experimental group and the control group were detected by RT-qPCR.In this study,ImageJ was used to analyze the gray value of protein bands,and SPSS 25.0 was used as the statistical analysis software.Results:1.Western blot showed that the expression of H3K27me3 in cholesteatoma of middle ear was lower than that in external ear canal.The statistical analysis showed that the mean ± standard deviation of the gray value ratio in the middle ear cholesteatoma was 0.72 ± 0.26,and the mean ± standard deviation of the gray value ratio in the outer ear canal was 1.40 ± 0.26.The difference was statistically significant(t=5.799,P<0.05).2.Immunofluorescence staining showed that H3K27me3 was mainly expressed in the nucleus of cholesteatoma cells in the middle ear and the outer auditory tract.After staining,the expression of H3K27me3 was red staining protein in the area of blue staining nucleus,and the expression of H3K27me3 in cholesteatoma cells was significantly lower than that in the outer auditory tract.3.RT-qPCR results showed that the relative expression of EZH2 mRNA in the experimental group decreased,t value was 5.20,P=0.0003,P<0.001,the difference was statistically significant,indicating that the expression level of EZH2 in the middle ear cholesteatoma was lower than that in the outer ear canal epithelium;the relative expression of EZH1 in the experimental group decreased,t value was 2.23,P=0.038,P<0.05,the difference was statistically significant,indicating that The expression level of EZH1 in middle ear cholesteatoma was lower than that in the epithelium of external auditory canal,but there was no significant difference in the expression of KDM6A,TGM1 and IVL.Conclusion:The expression of H3K27me3 in the middle ear cholesteatoma is lower than that in the outer ear canal,which may be involved in the formation and development of the middle ear cholesteatoma;H3K27me3 is mainly expressed in the nucleus of the middle ear cholesteatoma and the outer ear canal;EZH2 and EZH1 gene expression in the middle ear cholesteatoma is decreased,which may be the original factor affecting the down-regulation of H3K27me3 expression Therefore,it may provide a new target for the treatment of cholesteatoma of middle ear.