Ezrin Expression In Endometrial Carcinoma And Its Relations With Flt-1, TβRⅠ And TβRⅡ

Background Incidence rates of endometrial carcinoma have been increasing in recent years worldwide. Although gynecological oncologists at home and abroad have made a great deal of investigations, the molecular mechanisms that underlie the initiation and progression of endometrial carcinoma remain largely unknown, and the 5-year disease-free survival hasn't been elevated either. Ezrin is one of the membrane-cytoskeleton linking proteins. Many malignant tumors overexpress ezrin. Although the pathogenic mechanisms have been indicated to correlate with receptor tyrosine kinases such as HGF and EGF, it is yet unclear. Angiogenesis is vital to the growth and dissemination of solid tumors and VEGF is the most important angiogenic factor. VEGF receptors have traditionally been described as being expressed in endothelial cells;however, recent studies have demonstrated that VEGF receptors are also expressed in tumor cells. These findings suggest that circulating VEGF family members may not only promote tumor angiogenesis by activating VEGF receptors on endothelial cells but also have an autocrine effect on tumor cells bearing VEGF receptors. TGF-|3 signaling exerts a complex set of effects on cancers. Any disturbancesof TGF-β cascade components can allow the cells to evade the growth inhibitory effects of TGF-β. Some researchers discovered that dominant-negative ezrin inhibited the migration and clonogenicity of glioma cells and simultaneously induced a dramatic loss of TGF-β release. However, there has been no report on ezrin expression in endometrial carcinoma and its relations with flt-1, TβRI and TβRII. Expression of ezrin, flt-1, TβRI and TβRII in formalin-fixed, paraffin-embedded sections of normal endometrium, simple endometrial hyperplasia, complex endometrial hyperplasia, atypical endometrial hyperplasia and endometrial carcinoma was detected by immunohistochemistry and the association of these alterations with clinicopathological parameters in endometrial carcinoma was also evaluated. Furthermore, Western blot analysis was employed to measure ezrin expression in fresh-frozen tissues of endometrial carcinoma and normal endometrium.Objective To explore the role of ezrin in the initiation and progression of endometrial carcinoma and whether the underlying mechanism is corrrelated with TGF-p signaling and VEGF/flt-1 autocrine system or not.Material and methods 1. 140 tissue samples were obtained from the patients who underwent hysterectomy or diagnostic curettage between January 2002 and December 2003 at the affiliated Obstetrical and Gynecological Hospital, School of Medicine, Zhejiang University. The speimens of endometrial carcinoma were obtained from 79 patients(12 of them had metastatic lesions) with a median age of 55, and the specimens of atypical endometrial hyperplasia from 14 patients with a median age of 48, simple or complex endometrial hyperplasia from 20 patients with a median age of 45, normal endometrium from 15 patients with a median age of 45 were also obtained. 2. The clinicopathologic features of endometrial carcinoma such as age, FIGO stage, histological type, histological grade and myometrial invasion were all recorded. 3. Protein expression of ezrin, flt-1, TPRI and TpRII in all tissues was assessed by thePowerVision two-step immunohistochemical staining procedure. We observed all the slides with light microscope, identified the location of these four proteins, and made a semi-quantitative analysis with regard to the quantity and intensity of positively stained cells(H-score). Flesh tissues of endometrioid carcinoma from another 48 patients with a median age of 55 and normal endometrium from 48 patients with a median age of 46 who underwent hysterectomy because of hysteromyoma were also obtained. Western blot analysis was employed to measure ezrin protein in these two groups. 4. Statistical analysis was performed by SPSS 11.0 for windows. All tests were conducted at a significance level of a =0.05(two-tailed).Results Expression of ezrin protein: Immunohistochemistry showed that ezrin immunoreactivity was different in the four groups of proliferative endometrium, simple or complex endometrial hyperplasia, atypical endometrial hyperplasia and endometrial carcinoma(P=0.000). Ezrin expression was significantly higher in atypical endometrial hyperplasia than in simple or complex endometrial hyperplasia and ezrin expression was significantly higher in endometrial carcinoma than in atypical endometrial hyperplasia. Ezrin expression was either negative or polarized in most normal and hyperplasia endometrial cells, whereas it showed more diffuse staining of the cytoplasm and cell membrane in cancerous and atypical cells. Ezrin expression in endometrial carcinoma was statistically correlated with the depth of myometrial invasion(/M).014), and it was greater in papillary serous carcinoma and clear cell carcinoma than in endometrioid carcinoma(P=0.013). By Western blot analysis, ezrin protein was significantly greater in endometrial carcinoma than in normal endometrium(P=0.030). 2. Expression of flt-1 protein: Immunohistochemistry revealed that flt-1 expression in atypical endometrial hyperplasia was significantly higher than that in normal endometrium. However, flt-1 expression showed decreased intensity with increased tumor grade in endometrial carcinoma(P=0.018). 3. Expression of T|3RI and TPRII: Byimmunohistochemistry, TfJRI and TpRII were both lower in endometrial carcinoma than in normal endometrium, and the differences were statistically significant (P=0.011 and P=0.000, respectively). Furthermore, loss of TpRI and TpRII were both associated with deeper myometrial invasion (P=0.000 and P=0.008, respectively) and higher histologic grade (i^O.004 and P=0.000, respectively). 4. Correlation between ezrin and other cytokines such as fit-1,TpRI and TPRII: Ezrin was inversely correlated with TpRI in endometrial carcinoma^O.OOl), but it was not significantly correlated with flt-1 or TpRII.Conclusions 1. Ezrin overexpression and defective expression of TGF-p receptors were both involved in the carcinogenesis and progression of endometrial carcinoma and the correlation between ezrin and TGF-p receptor was significant, suggesting that they might coparticipate in the carcinogenesis and progession of endometrial carcinoma through interacting with each other. 2.The fact that ezrin expression was related to histologic type and myometrial invasion implies that ezrin may be associated with the prognosis of endometrial carcinoma patients. 3. The role of ezrin in endometrial carcinoma is not related to VEGF/flt-1 autocrine pathway. VEGF/flt-1 autocrine pathway may merely contribute to the early endometrial malignant transformation.