| Background and purpose:Ovarian cancer is one of the most lethal tumors of the female genital tract.The 5-year survival rate for advanced epithelial ovarian cancer is less than 50%.The high mortality rate of ovarian cancer is associated with its invasiveness and metastasis potential,but the underlying mechanism is not clear.Mediator is a large multisubunit complex involved in transcription initiation.Previous studies have found that the mediator complex 29 subunit(MED29)is highly expressed in ovarian cancer tissues and cells,which can promote the proliferation of ovarian cancer cells.However,the effects of MED29 on the invasion and migration abilities of ovarian cancer cells and its mechanism have not been deeply discussed.Therefore,this study aims to explore the effects of MED29 on the invasion and migration abilities of ovarian cancer cells by combining clinical data and cell experiments and to further verify the molecular mechanism by using RNA sequencing technology,so as to provide a new target for ovarian cancer diagnosis and treatment.Methods:1.The bioinformatics analysis: The R software was used to analyze the ovarian cancer data in the GEO database and the TCGA database,and MED29 expression levels between normal ovarian or fallopian tube tissues and ovarian cancer tissues were compared.2.The clinical data analysis: Immunohistochemical analysis was performed on tissue microarrays from 37 patients(including 32 epithelial ovarian cancer tissues and5 normal ovarian tubal tissues)to determine the relationship between MED29 expression level and ovarian cancer metastasis.3.Cell phenotypic experiment: Two small interfering RNA(si RNA)sequences targeting MED29 were transfected into ovarian cancer cell lines(A2780 and HEY)to construct the MED29 knockdown cell model.The wound healing assay and the Transwell cell invasion and migration assay were used to observe the effect of MED29 on the ovarian cancer cell phenotype.In addition,the expression levels of EMT-related proteins were examined by Western blot.4.Regulatory analysis of MED29 on downstream molecules: The si RNA sequence with a better knockdown effect was selected to transfect A2780 cells.Transcriptome sequencing was performed to analyze the regulatory effect of MED29 on downstream molecules at the transcription level.The downstream signaling pathway of MED29 involved in the invasion and migration of ovarian cancer cells was determined,and the expression of this pathway was verified.Results:1.MED29 was much more expressed in ovarian cancer tissues than in healthy ovarian or fallopian tube tissues,according to the findings of a bioinformatics investigation.2.Immunohistochemistry analysis revealed that the expression of MED29 was significantly higher in ovarian cancer tissues than in healthy ovarian tubal tissues.When compared to ovarian cancer tissues without metastasis,the expression of MED29 was considerably higher in tissues with pelvic invasion and/or abdominal metastasis.The expression of MED29 was positively correlated with TNM stage and metastasis,but not with age.3.Ovarian cancer cells transfected with si RNA considerably decreased their ability to invade and migrate when compared to the negative control group,according to cell phenotypic data.These findings were statistically significant(P<0.05).Western blot results showed that the expression levels of EMT-related proteins,N-cadherin and Vimentin,were significantly inhibited,while the expression of E-cadherin was significantly up-regulated after interfering with MED29.4.KEGG enrichment analysis showed that MED29 may regulate the invasion and migration of ovarian cancer cells through the ERK signaling pathway.Western blot results showed that p-ERK/ERK expression levels were significantly decreased in ovarian cancer cells after interference with si RNA-MED29(P<0.05).Conclusions:1.MED29 is highly expressed in ovarian cancer tissues and is positively correlated with ovarian cancer metastasis and TNM stage.2.MED29 can promote the migration and invasion of ovarian cancer cells by regulating the ERK1/2 pathway to mediate EMT. |
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