Estradiol Regulates NF-κB Activity Via AKT Signaling Pathway In Endometrial Cancer

Background Endometrial cancer is common gynecologic malignancy, that the morbidity is second to cervical cancer in China. With the average life expectancy of women and, estradiol replacement therapy increased, the addition in the proportion of obesity and other factors, the incidence of endometrial cancer in all age groups is rising both in western countries and china in recent years, but its reason is not very clear. It may be related to over-exposure to estradiol that is not inhibited by progesterone.AKT (Serine/threonine protein kinase) is closely related to cell survival and apoptosis,is downstream target of PI3k (Phosphoinositide 3-kinase). PI3k/Akt is a classical signaling pathway, playing an important role in signal transduction, is the cross-piont of various pathways.NF-κB (nuclear factor-kappa B) is an important nuclear transcription factor. Its protein family is usually bind to DNA sites as dimers. The physiological function of heterodimer is composed of p50 and p65/Re1A. When cells are stimulated, endogenous inhibitory protein IκB (inhibitory protein of NF-κB) degradation, activited NF-κB is released and move to the nucleus to regulate gene transcription. NF-κB signaling pathway is relate to the tumor cellular processes, such as occurrence proliferation, differentiation, apoptosis, invasion and metastasis. More and more evidences show that NF-κB is a powerful tumor-promoting factor, and It has become a new target for cancer treatment.Objective The main aims are to study if NF-κB can be activated by estradiol via AKT signaling pathways. In this study, Endometrial cancer cell lines HEC-lA(ER-weak positive) and Ishikawa (ER-positive) were treated by estradiol, then examined the NF-KB activity to described the relationship between NF-κB and estradiol in endometrial carcinoma.Method1,Western Blot was detected the expression of AKT protein in human endometrial cancer cell lines HEC-1A,Ishikawa after stimulation with 1×10-6mol/L,tradiol for 30min; and with being pretreated with 25x 10-6mol/L AKT inhibitor (AKT inhibitor group)or 1×10-6mol/L ER inhibitor(ER inhibitor group) for 60min, following stimulation with 1×10-6mol/L estradiol for 30min,2,Used the TransAMTM NFκB p65 Kit to detect the NF-κB p65 activity in HEC-1A,Ishikawa cells. The cell lines were incubated with increasing concentrations of estradiol (10-4mol/L.10-6mol/L,10-8mol/L,10-10mol/L) for 30min; or the cell lines were exposed to E2 (1×10-6mol/L) for specific time intervals (15,30,1h and 2h);or the cell lines were pretreated with the 25×10-6mol/L AKT inhibitor for 1h, following stimulation with 1×10-6mol/L estradiol for 30min (AKT inhibitor group).Result1,Western Blot showed that the expression of p-AKT protein in HEC-1A,Ishikawa cells after stimulation with 1×10-6mol/L estradiol for 30min was markedly higher than that in the control group (P﹤0.05); the expression of p-AKT protein expression in AKT inhibitor group and ER inhibitor group were significantly decreased than that in estradiol group(P<0.05).2,Dose-dependent activation of NF-κB by estradiol:The NF-κB activity in HEC-1A,Ishikawa cell lines after stimulation with 1×10-6mol/L estradiol were significantly higher than the control group and the other concentrations groups (P<0.05);time-dependent activation of NF-κB by estradiol:1×10-6mol/L estradiol induced a significant increase of NF-kB activity in HEC-1A,Ishikawa cell lines at 30min and 1h, compared with 15min and 2h (P﹤0.05),It showed a relatively increased between 30min and 1h but no significant difference; the NF-κB activity of AKT inhibitor group were significantly reduced compared to 1×10-6mol/L estradiol group (P<0.05).Conclusion Estradiol can regulate the activity of NF-κB, may be through the AKT signaling pathway.