The Expression And Significance Of IKKε In The Specimens And Cells Of Human Endometrial Carcinoma

Objective:To detect the protein expression of IKKε in the specimens and cell lines of human endometrial carcinoma,and meanwhile the impact that IKKε inhibitor has on cell proliferation,apoptosis as well as IKKε expression of endometrial carcinoma cells.To explore the role of IKKε in human endometrial carcinoma.Methods:(1)Immunohistochemistry staining method was employed to determine the protein expression of IKKε in various specimens.The specimens were taken from 107 patients who had a diagnosis of endometrial carcinoma and received the hysterectomy from January of 2011 to September of 2014.33 cases with endometrial carcinoma was randomly selected,from which 33 cases of peritumoral normal endometrium were also taken correspondingly as the control group.Meanwhile,we did the follow-up visit to those 107 patients,and the correaltionship of IKKε expression and different clinicopathological factors of endometrial carcinoma was also analyzed.(2)Human endometrial cancer cell lines Ishikawa(derived from well differentiated endometrial cancer),HEC-1A,HEC-1B(both derived from moderately differentiated endometrial cancer)and AN3CA(derived from undifferentiated endometrial cancer)were incubated in vitro.Western blot was applied to detect IKKε expression in all of those four cell lines and the effect of IKKε inhibitor A(Malachite Green Oxalate)(0.1μM、0.5μM)on the protein expression.The cell proliferation was detected by methyl thiazolyl tetrazolium(MTT)method treated with IKKε inhibitor A with different concentrations(0-25μM).The apoptosis rate after 48 hour-treatment of IKKε inhibitor A with different concentrations(0μM,0.1μM,0.5μM)was analyzed by flow cytometry.Results:(1)IKKε protein is mainly expressed in cytoplasm.The positive expression rate of IKKε in peritumoral normal endometrium and endometrial carcinoma were 51.5% and 72.7%,respectively.Apparently,IKKε expression in endometrial carcinoma was greater than it in the control group,though not significantly(P=0.076>0.05).Another findings of ours is that the expression of IKKε had no correlation with the clinicopathological factors such as age,FIGO classification,pathological grade,pathological type,muscular invasion depth,the level of serum CA125,survival duration or complications including hypertension and diabetes mellitus(In all cases,P>0.05).(2)IKKε protein expression is observed in all the four cell lines: Ishikawa、HEC-1-A、HEC-1-B and AN3 CA.After treating the four cell lines with IKKε inhibitor A(0μM,0.1μM,0.5μM)for 48 hours,we noticed that the protein expression decreased.(3)MTT assays indicated that the cell proliferation was effectively inhibited after 48 hours of IKKε inhibitor A with different concentrations(0-25μM)(P<0.05).IC50 of Ishikawa、HEC-1A,HEC-1B and AN3 CA were 0.134μM,0.431μM,1.073μM and 0.079μM,respectively.(4)The apoptotic rates increased after 48 hour-treatment of IKKε inhibitor A with different concentrations(0μM,0.1μM,0.5μM),and the result varied among the four cell lines.Importantly,Ishikawa and AN3 CA ended up with significant cell apoptosis after the inhibitor,while the same phenomenon was not yet able to be observed in HEC-1A and HEC-1B(P>0.05).Conclusions:IKKε is highly expressed in endometrial carcinoma.And in vitro,IKKε protein expression at different levels is also detectable in different endometrial cancer cell lines.The moderately differentiated endometrial cancer cells have significantly higher IKKε expression than the well differentiated Ishikawa cells and undifferentiated AN3 CA cells.IKKε inhibitor A,downregulating the protein expression,may surppress cell proliferation and contributes to cell apoptosis of endometrial cancer cells.The increasing concentration of the inhibitor results in stronger surpression in cell proliferation,and the well differentiated and undifferentiated endometrial carcinoma cells have an improved sensitivity to IKKε inhibitor A compared with the moderately differentiated ones.Hence,we conclude that IKKε might play an important role in the survival,prolifereation and anti-apoptosis of endometrial cancer cells.We also speculate that IKKε might be involved in the development and progress of endometrial carcinoma,and will promisingly become a novel option for the molecular targeted therapies of endometrial carcinoma.