Relationship Between SOX8 And Clinicopathology Of Endometrial Cancer Patients And Its Influence On Cell Biological Behavior

Objective: Endometrial cancer is one of the three major malignant tumors that threaten women’s reproductive health,and its incidence is increasing year by year.However,the mechanism of endometrial cancer remains unclear.The purpose of this study is to understand the role of SOX8 gene in occurrence and development of endometrial cancer,to analyze the relationship between SOX8 and clinicopathological features and prognosis of endometrial cancer patients.Further,we want to explore the effect and mechanism of SOX8 gene on the proliferation,migration and invasion of endometrial cancer cells.Method: 1.Clinical data of 274 patients with endometrial cancer and 160 normal endometrium,116 proliferative endometrium,and 76 atypical endometrial hyperplasia in the same period was collected.Immunohistochemistry was conducted to explore the expression of SOX8 in four types of endometrium tissues,to analysis the correlation between the expression of SOX8 and the clinicopathological characteristics and prognosis of EC patients.2.Western blot method was used to detect the basic level of SOX8 in ECC-1,Ishikawa,HEC-l A,HEC-l B,and KLE.Ishikawa and HEC-l B cell lines were used for overexpression and know-down of SOX8 by lentivirus transfection and RNA interference experiments,respectively.Western blot was used to verify the effects of up or down regulation.CCK8 and Clonal formation methods were used to detect the proliferation ability of EC cells.Transwell assay was used to detect the migration and invasion ability of EC cells.Results: 1.We examined the expression of SOX8 in NE,HE,AHE,and EC groups on TMAs via immunohistochemistry.The expression of SOX8 in EC tissues was higher than that in NE tissues(χ2=11.685,P=0.003).Among the 277 EC patients,high expression of SOX8 was significantly associated with high tumor histological grade(χ2=6.907,P=0.032)and high rate of lymph node metastasis(Fisher,P=0.027).The expression level of SOX8 was higher in deep myometrial invasion,advanced FIGO stage and 2 type EC samples,though did not reach a statistic significance.(P>0.05).Results from Kaplan-Meier curve show that patients with high SOX8 expression was associated with shorter OS when compared to patients with low SOX8 expression and negative expression(χ2=6.930,P=0.031).2.Five commonly EC cell lines were used to examine endogenous SOX8 expression by western blot.SOX8 protein expression was relatively higher in HEC-1B cells,lower in Ishikawa cells.HEC-1B and Ishikawa cells were chosen for the following research.We established SOX8-knockdown cells and SOX8-overexpressing cells to determine whether there were alterations in cell proliferation,migration,and invasion.CCK8 assay demonstrated that the cell growth rates in SOX8 knockdown HEC-1B group were significantly decreased compared with control and NC group(P<0.05),Colony formation assays demonstrated that the colony number of SOX8 knocking down group was significantly decreased compared to control and NC group(251.3 ± 23.25,436.3 ± 9.615 vs 732 ± 14.57,vs 744.3 ± 32.2,P<0.001),transwell assay shows that knockdown of SOX8 in HEC-1B cells inhibits cell migration compared to control and NC group(17.4±1.364,22.8±1.393 vs 70±3.619,vs 73±2.387,P<0.0001)also as invasion assay(7±0.3162,15.2±0.860 vs 51.8±1.685,vs 50.6±2.4,P<0.0001).3.Conversely,CCK8 assay shows that overexpression of SOX8 significantly promoted the proliferation in Ishikawa cells(P<0.01).Colony formation assays demonstrated that over expression of SOX8 could increase the number of foci compared to control and NC group(681 ± 20 vs 427 ± 1,vs 403 ± 5,P<0.01).transwell assay shows that overexpression of SOX8 enhances cell migration(81.4±5.437 vs 26.2±1.02,vs 31.8±2.99,P<0.0001)and invasion(66.2±2.709 vs 21±0.8367,vs 19.4±2.694,P<0.0001)ability compared to control and NC group.Conclusion: 1.The expression of SOX8 was gradually increased from NE,HE,AHE to EC tissue,suggesting that SOX8 may be related to the occurrence of endometrial cancer.2.High expression of SOX8 was associated with poor differentiation and positive lymph node metastasis.The prognosis of patients with high expression of SOX8 is poor,suggesting that SOX8 may participate in the malignant evolution of endometrial tissue.SOX8 may be a molecule that indicates the prognosis of EC patients.3.SOX8 can promote the proliferation,migration and invasion of EC cells,suggesting that SOX8 may be an oncogene for EC.