| Prat 1.The Expression Profile and Clinical Significance of MAP3K3 in Ovarian CarcinomaObjective:To investigate the expression profile of MAP3K3 in Ovarian carcinoma(OC)and to assess its values in clinical biological behabior and prognosis of OC patients.Methods:Immunohistochemistry staining and quantitative RT-PCR testing for MAP3K3 expression was performed on 93 primary ovarian tumors and 33 normal fallopian tube tissue samples,and the correlation between MAP3K3 expression with clinicopathological factors of OC were investigated further.Results:(1)MAP3K3-positive staining predominantly showed cytoplasmic localization,and all OC specimens were positive for MAP3K3 expression.It markedly varied between OC and normal fallopian tube tissue.MAP3K3 overexpression was detected in 55 of 93(59.1%)cases of OC and 10 of 33(30.3%)cases of fallopian tube,indicating a significant difference(p<0.05).(2)The expression level of MAP3K3 mRNA in OC was significantly higher than that in normal fallopian tube tissue(p<0.01).(3)MAP3K3 overexpression was significantly associated with the histological type(p = 0.001),histological grade(p = 0.018),challenge model(p = 0.000),and chemotherapy response(p = 0.009).No significant correlations were observed between MAP3K3 expression and other clinicopathologic parameters,including patient age(<50 years vs.>50 years)(p = 0.434),FIGO stage(p=0.491),and ascites(p = 0.988).(4)Kaplan-Meier survival analysis indicated that patients with MAP3K3 overexpression exhibited a significantly reduced DFS and OS than those with low expression(log-rank test,p = 0.007 and p = 0.001,respectively);In a univariate analysis based on the Cox regression model,MAP3K3 overexpression was considered a strong prognostic factor of poor OS(p= 0.036;Table 3).In addition,age(p = 0.031),chemotherapy response(p = 0.003),and FIGO stage(p = 0.007)were associated with shorter OS.The multivariate COX analysis identified MAP3K3 overexpression(HR:2.082,95%CI=1.154-3.756,P= 0.015),age(HR:4.127,95%CI=1.178-14.461,p = 0.027),chemotherapy response(HR:3.299,95%CI= 1.473-7.387,P= 0.004),and FIGO stage(HR:2.038,95%CI=1.083-3.834,p = 0.026)as independent prognostic factors,whereas histological type(p = 0.380)and grade(p = 0.328),challenge model(p = 0.651),and ascites(p = 0.371)were identified as insignificant independent prognostic factors.Conclusion:(1)MAP3K3 protein and mRNA are highly expressed in ovarian carcinoma tissues,and MAP3K3 protein expression is associated with histopathological grade,pathogenesis and chemotherapy response of ovarian cancer.(2)Tumor-free survival and the overall survival time of patients with high expression of MAP3K3 protein was significantly lower,the high expression of MAP3K3 protein is an independent risk factor for poor prognosis in ovarian carcinoma.Part 2 The Effect of MAP3K3 Mediated NF-κB Signaling Pathway on The Biological Behavior of Ovarian CarcinomaObjective:To investigate the effects of MAP3K3 on the malignant biological function of ovarian carcinoma,such as proliferation,apoptosis,invasion and metastasis,and to explore the possible pathway of MAP3K3 gene mediated in the development of ovarian carcinoma.Methods:In OC cell lines,MAP3K3 expression was evaluated by Western blot analysis,quantitative RT-PCR,and immunofluorescence.MAP3K3 eukaryotic expression vector and MAP3K3 specific interference expression vector were used to transfect ovarian cancer cell lines with low expression and/or high expression of MAP3K3.The transfection efficiency was verified by Western Blot and qRT-PCR.The expression of NF-κB signaling pathway and the expression of EMT-related factors were observed before and after treatment The active state of NF-κB signal pathway and the expression of EMT-related factors were observed before and after treatment with NF-κB signal-specific blocker QNZ.CCK-8 was used to detect the proliferation of ovarian carcinoma cells before and after the interference.The colony formation assay was used to detect cell clone formation before and after interference.Flow cytometry was used to detect apoptosis,scratch experiments and Transwell chamber experiments to detect migration and invasion.EMT-related proteins and cytoskeletal proteins were detected by immunofluorescence.Results:(1)During 6 OC cell lines,High MAP3K3 expression was significantly detected in SKOV3,C13*,and A2780 cells,whereas reduced expression was detected in HeyA8,OVCA433,and OV2008 cells.(2)The down-regulation of MAP3K3 could inhibit the activation effect of TNF-α on NF-κB signaling pathway,the activity of NF-κB signal pathway decreased after MAP3K3 knockdown,and the expression of the mesenchymal markers N-cadherin,Vimentin and ICAM1 were down-regulated,Epithelial marker E-cadherin expression was up-regulated,while overexpression of MAP3K3 could revers those above phenomenon.(3)Celluer function experiments showed that knockdown of MAP3K3 could inhibit the proliferation,colony formation,migration and invasion of ovarian carcinoma cells,and lead to the increase of spontaneous apoptosis in vitro,while overexpression of MAP3K3 could reverse the above phenomenon.When overexpress MAP3K3 on the basis of blocking NF-κB signaling pathway,proliferation,colony formation,migration and invasion ability of ovarian carcinoma cells decreased,spontaneous cell apoptosis increased.Conclusion:MAP3K3 gene can promote the proliferation and invasion of ovarian cancer cells,and could enhance the migration and invasion of ovarian carcinoma cells.MAP3K3 gene may regulate ovarian carcinoma malignant biological behevior by mediating NF-κB signaling pathway,through affecting epithelial mesenchymal transition and cytoskeletal protein expression. |
PDF Full Text Request