The Preliminary Study Of The Functional Role Of RAC1 In The Migration And Invasion Of Endometrial Cancer And Its Underlying Mechanism

Endometrial cancer(EC)is a malignant tumor originating from the endometrium,which is the second most common malignant tumor of the female reproductive system in China.Statistics show that the 5-year survival rate of endometrial cancer has not improved significantly in 30 years.The tissue invasiveness and distant metastatic ability of EC seriously affect the prognosis.The metastatic mechanism of EC has not been elucidated.This study found that RAC1 gene was closely related to EC by gene chip and bioinformatics,and its expression in EC tissue was higher than that in normal endometrial tissue.RAC1 may promote the invasion and migration abilities of EC by increasing the number of lamellipodia.This effect may be related to the regulation of CTTN by RAC1.Chapter 1 Differentially expressed genes profiling in endometrial cancerObjective To explore key differentially expressed genes(DEGs)and pathways in endometrial cancer for a better understanding of molecular mechanism.Methods Microarray was carried out on 3 fresh endometrial cancer(EC)specimens and 3 fresh normal endometrium specimens.Bioinformatics was used to analyze the DEGs,significant enrichment of pathways,disease and function,and construction of Protein-Protein Interaction Networks.Finally,GEPIA and UALCAN analyzed the expression of RAC1 in EC and normal endometrium.Results A total of 916 DEGs were screened of which 480 genes were up-regulated and 436 were down-regulated.The results showed that the significant functional process included cell development,cell growth and proliferation,cancer,cell assembly and organization,cell movement and so on.The significant signal pathways involved in up-regulated DEGs were: P53Signaling、ERK/MAPK Signaling、PTEN Signaling and so on.It was found that RAC1 is one of the key up-regulated DEGs and participates in the key signal pathway of tumor.There was no significant difference in the expression of RAC1 m RNA in EC tissue(p>0.01),but there was an increasing trend,while the protein expression of RAC1 was significantly increased in EC tissue,and the difference was statistically significant(p<0.001).Conclusion It is found that RAC1 is one of the key up-regulated deferentially expressed genes in endometrial cancer and participates in the key signal pathway of tumor by microarray incorporated with bioinformatics.Chapter 2 Expression of Ras-related C3 botulinum toxin substrate 1 and Cortactin and in endometrial cancer and its clinical significanceObjective To investigate the expression of Ras-related C3 botulinum toxin substrate 1(RAC1)gene and Cortactin(CTTN)gene in endometrial cancer tissue and its clinicopathologic features.Methods The clinical specimens of endometrial cancer and normal endometrium and the clinical data of corresponding patients were collected.Total RNA were extracted,and tissue paraffin sections were prepared.Real-time Quantitative PCR(q PCR)was used to detect the relative expression level of RAC1 m RNA and CTTN m RNA in endometrial cancer tissue and normal endometrial tissue.Immunohistochemical staining(En Vision method)was used to detect the protein expression of RAC1 and CTTN in endometrial cancer tissue and normal endometrial tissue.The relationship between the expression of the RAC1 gene and the CTTN gene and the prognostic factors was analyzed.Results The m RNA relative expression level and protein expression level of RAC1 and CTTN and in endometrial cancer group were significantly higher than those in normal endometrial group(p<0.05),and there was correlation between the m RNA and protein expression of RAC1 and CTTN.The protein expression of RAC1 was not related to pelvic lymph node invasion or histologic grade,but was related to lymph-vascular space invasion(LVSI),deeper myometrial invasion and FIGO stage(p < 0.05).The protein expression of CTTN was not related to pelvic lymph node invasion,LVSI or histologic grade,but was related to deeper myometrial invasion and FIGO stage(p<0.05).Conclusion The RAC1 gene and CTTN gene may play a role in the occurrence and development of endometrial cancer,and there is a correlation between the two genes.Chapter 3 The Study of effect of RAC1 on migration and invasion of endometrial cancer cells and its mechanismObjective To explore the effect of RAC1 on the invasion and migration of endometrial cancer cells and its potential mechanism.Methods lentiviral system was constructed and transfected into HEC-1-A and HEC-1-B cells.The migration and invasion abilities of the cells in each group were evaluated by scratch assay and transwell migration and invasion assays.Pseudopodia formation was observed using immunofluorescence staining.Western blot and q PCR were used to detect RAC1 and CTTN expression.Results After CTTN knockdown,the migration rate,invasion ability,and migration ability of HEC-1-A and HEC-1-B cells decreased significantly.A decrease in the number of lamellipodia,accompanied by a compensatory increase in the number of blebs,was also observed.Overexpression of RAC1 promoted the migration and invasion of HEC-1-A and HEC-1-B cells(p < 0.05),and the effect of CTTN knockdown on cell migration and invasion ability could be reversed.Knockdown of CTTN had no significant effect on the expression of RAC1 but the expression of CTTN increased after overexpression of RAC1.The expression of RAC1 increased after cotransfection,but there is no significant effect on the expression of CTTN.Conclusion RAC1 may promote the invasion and migration abilities of EC by increasing the number of lamellipodia.This effect may be related to the regulation of CTTN by Rac1.