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E-Cadherin Expression in Endometrial Malignancies: Comparison between Endometrioid and Non-Endometrioid Carcinomas T Yalta, L Atay, F Atalay, M Çaydere, M Gonultas and H Ustun Journal of International Medical Research 2009 37: 163 DOI: 10.1177/147323000903700119 The online version of this article can be found at: http://imr.sagepub.com/content/37/1/163
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The Journal of International Medical Research 2009; 37: 163 – 168 [first published online as 37(1) 1]
E-Cadherin Expression in Endometrial Malignancies: Comparison between Endometrioid and Non-endometrioid Carcinomas T YALTA1, L ATAY2, F ATALAY2, M ÇAYDERE2, M GONULTAS2
AND
H USTUN2
1
Pathology Department, Sivas State Hospital, Sivas, Turkey; 2Pathology Department, Ankara Training and Research Hospital, Ankara, Turkey
This study examined the frequency of Ecadherin expression in endometrial biopsy or hysterectomy specimens from patients diagnosed with endometrial adenocarcinoma and in normal endometrial tissue specimens. E-cadherin expression was detected by immunohistochemistry using monoclonal antibody to E-cadherin. Specimens were classified as positive when ≥ 5% of the tumour cells showed staining for E-cadherin, irrespective of the pattern of staining. Twenty-three endometrioid carcinomas and nine non-endometrioid (four papillary serous and five clear cell) carcinomas were studied, along with 10 normal endometrial tissue specimens as KEY WORDS: E-CADHERIN; ENDOMETRIOID
Introduction Among the female genital tract cancers, endometrial carcinoma is the most common one in most European countries,1 but overall survival is relatively higher compared with other gynaecological malignancies.2 Patients with non-endometrioid histology (papillary serous or clear cell), tumoural invasion deep into the myometrium, poor tumour differentiation, or the presence of metastasis
controls. E-cadherin expression was significantly less frequent in nonendometrioid carcinomas compared with endometrioid carcinomas and controls. There was no statistically significant difference in the frequency of E-cadherin expression between endometrioid carcinomas and controls. In conclusion, this study demonstrated that uterine nonendometrioid (papillary serous and clear cell) carcinomas were less likely to express E-cadherin compared with endometrioid carcinomas and normal endometrial tissue. This may help to explain the more aggressive behaviour of non-endometrioid carcinomas. CARCINOMA;
NON-ENDOMETRIOID
CARCINOMA
(to other organs or lymph nodes within the pelvis) have a worse prognosis and are at increased risk for recurrence after treatment.3 Previous studies have suggested that poor survival in non-endometrioid carcinomas is the result of their increased tendency for extra-uterine metastasis.4 This invasive behaviour contrasts with that of endometrioid carcinomas, which are generally confined to the uterus at the time
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T Yalta, L Atay, F Atalay et al. E-cadherin expression in endometrial carcinoma of diagnosis. It has been propounded that the aggressive behaviour of some carcinomas might be due to decreased intercellular cohesiveness in these tumours. Cell-to-cell adhesion is mediated by transmembrane glycoproteins known as cadherins. These are divided into subgroups on the basis of their tissue distribution: E-cadherin (epithelial), Pcadherin (placental), N-cadherin (neural) and L-cell adhesion molecule (liver). E-cadherin has been shown to play a central role in the organization and maintenance of epithelial tissue structure. Decreased cell-to-cell contact in epithelial cells has been shown to be largely attributable to down-regulation in the expression of E-cadherin.4 In vitro studies have demonstrated that expression of Ecadherin may preclude the invasiveness of epithelial tumour cell lines, suggesting that the E-cadherin gene may primarily act as an invasion-suppressor gene.5 Canine kidney epithelial cells were found to acquire more invasive properties when E-cadherin was specifically blocked by anti-E-cadherin antibodies, supporting the fact that decreased E-cadherin expression is a crucial step in the progression to a more malignant phenotype.6 Decreased E-cadherin expression has also been associated with decreased cell-to-cell adhesion and increased invasive and metastatic potential in endometrial and other carcinomas.7 The present study was devised to examine and compare E-cadherin expression in endometrioid and non-endometrioid (papillary serous and clear cell) endometrial carcinomas and normal endometrial tissue.
Materials and methods SOURCE OF SPECIMENS Endometrial biopsy or hysterectomy specimens from patients diagnosed with
endometrial adenocarcinoma and also normal endometrial tissue specimens were examined. All tissue samples were formalinfixed, paraffin-embedded and obtained from the tissue bank of the Pathology Department of Ankara Training and Research Hospital. A section from each specimen block was stained with haematoxylin and eosin for histological evaluation, and representative blocks were used for immunohistochemical studies.
IMMUNOHISTOCHEMICAL STAINING Specimens from the paraffin-embedded block were cut into 4 µm sections. They were then dewaxed using xylene and transferred to alcohol. Antigen retrieval was performed by microwaving for 15 min in 10 mM citrate buffer pH 6. Endogenous peroxidase was blocked by immersion in hydrogen peroxide for 10 min at room temperature. After washing in 25 mM phosphate-buffered saline (PBS) pH 7.6, the sections were incubated with ultraviolet block for 5 min. After further washing with PBS, each section was incubated with one drop (150 µl) of primary antibody (E-cadherin antimouse monoclonal antibody; NeoMarker, Fremont, CA, USA) for 60 min, washed with PBS and then incubated with biotinylated goat antipolyvalent antibody (ScyTek Laboratories, Logan UT, USA) for 10 min. After incubation of each specimen with 150 µl streptavidin peroxidase (ScyTek) for 10 min and washing with PBS, each specimen was reacted with 10 µl of 70 µl/ml diaminobenzidine chromogen solution, counterstained with haematoxylin and dehydrated in alcohol. A minimum of 25 fields were examined at ×10 magnification for each specimen. In general, there are four different staining patterns for E-cadherin: diffuse linear, when
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T Yalta, L Atay, F Atalay et al. E-cadherin expression in endometrial carcinoma crisp membrane staining is seen in > 75% of tumour cells in the absence of cytoplasmic staining; diffuse granular, when membrane and cytoplasmic staining is seen in 26 – 100% of the tumour cells; and focal granular, when membrane and cytoplasmic staining is seen in 5 – 25% of the tumour cells. In this study, specimens were classified as positive when ≥ 5% of the tumour cells showed staining for E-cadherin and as negative when < 5% of the tumour cells showed staining, irrespective of pattern.
STATISTICAL ANALYSIS Statistical analysis of the relationship between endometrial carcinoma type and Ecadherin staining was performed using Fisher’s exact test. A P-value < 0.05 was considered to be statistically significant.
Results A total of 32 endometrial biopsy or hysterectomy specimens from patients with endometrial adenocarcinoma were examined. Of these, 23 were endometrioid carcinomas, while nine were nonendometrioid carcinomas (four serous papillary, five clear cell). They were compared with a control group comprising 10 normal endometrial tissue specimens. Table 1 shows the number of E-cadherinpositive and -negative patients in each histological group. Non-endometrioid (papillary serous and clear cell) carcinomas were significantly less likely to express Ecadherin than endometrioid carcinomas or controls (P < 0.05). The difference in Ecadherin expression between endometrioid carcinomas and the control group was not
TABLE 1: E-cadherin expression n (%) in endometrioid and non-endometrioid (papillary serous or clear cell) endometrial carcinomas E-cadherin expression Type of carcinoma Endometrioid (n = 23) Non-endometrioid (n = 9) Control (n = 10)
Positive
Negative
18 (78.3%) 2 (22.2%)a,b 8 (80.0%)
5 (21.7%) 7 (77.8%) 2 (20.0%)
a
P < 0.05 versus endometrioid carcinoma. P < 0.05 versus control.
b
A
B
FIGURE 1: (A) and (B) Microscopic appearance of two different E-cadherin-positive endometrioid carcinomas (original magnification ×100; immunohistochemical staining with E-cadherin specific mouse monoclonal antibody)
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T Yalta, L Atay, F Atalay et al. E-cadherin expression in endometrial carcinoma
Apoptosis (cell death) is a tightly regulated cell cycle process. Expression of E-cadherin is not only critical for the regulation of intercellular cohesiveness, but also for the regulation of the apoptosis of tumour cells.8 Tumour growth is associated with both proliferation and apoptosis; the balance between these processes determines the number of tumour cells that are present.8 E-cadherin is a calcium-dependent
epithelial adhesion molecule.9 Loss of Ecadherin expression has been encountered in various types of human malignancies.10 A study on high grade neuroepithelial tumours demonstrated that loss of e-cadherin expression was associated with dedifferentiation and dissemination in these tumours.10 Cadherins are the primary molecules that play a central role in the maintenance of cell – cell junctions through a calcium-dependent cellular mechanism.10 In many malignancies, decreased E-cadherin expression is associated with defective cell – cell adhesiveness, resulting in invasion and metastasis.11 E-cadherin may, therefore, be considered to be an invasion-inhibiting
FIGURE 2: Microscopic appearance of an E-cadherin-negative clear cell endometrial carcinoma (original magnification ×100; immunohistochemical staining with E-cadherin specific mouse monoclonal antibody)
FIGURE 3: Microscopic appearance of an E-cadherin-negative papillary serous endometrial carcinoma (original magnification ×100; immunohistochemical staining with E-cadherin specific mouse monoclonal antibody)
statistically significant. The microscopic appearances of the endometrial carcinomas and the control group are shown in Figs 1 – 4.
Discussion
A
B
FIGURE 4: (A) and (B) Microscopic appearance of two different E-cadherin-positive normal endometrial tissue specimens (original magnification ×100; immunohistochemical staining with E-cadherin specific mouse monoclonal antibody)
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T Yalta, L Atay, F Atalay et al. E-cadherin expression in endometrial carcinoma molecule. In contrast, however, Shimoyama et al.12 found a human lung cancer cell line that did not demonstrate tight cell–cell adhesion yet was shown to be positive for E-cadherin. It was suggested that these cells were deprived of α-catenin (a cadherin-related molecule), leading to dysfunctional E-cadherin and decreased cell–cell adhesiveness. Patients with non-endometrioid carcinomas (papillary serous and clear cell) are more likely to have positive peritoneal cytology, lymph node metastasis and adnexal involvement, and lymph node and lymphovascular invasion than those with endometrioid tumours. For example, Goff et al.13 detected extra-uterine metastases in 36 out of 50 patients with uterine papillary serous tumours; myometrial invasion was not significantly associated with the extent of extra-uterine disease. Hoffman et al.14 suggested age as a significant risk factor, reporting that older patients had more nonendometrioid and histologically less differentiated endometrial cancers. Some previous studies, for example Furuyama et al.15 have drawn attention to the presence or absence of E-cadherin expression in cancers, reporting that loss of E-cadherin expression is significantly associated with peritoneal dissemination of tumour cells, leading to positive peritoneal cytology. Miyamoto et al.16 examined 30 well differentiated and poorly differentiated endometrioid carcinomas in terms of the expression and location of E-cadherin and some E-cadherin-related cytoplasmic
molecules, including α-catenin, β-catenin, γcatenin, p120CAS and IQGAP1 (molecules known to regulate the function of Ecadherin). They reported significant differences in the patterns of expression and location of these molecules between the two histological types of endometrioid carcinomas, suggesting that alterations in Ecadherin-related molecules might play different roles in E-cadherin dysfunction in tumours with different degrees of differentiation. In general, the frequency of E-cadherin expression in endometrioid carcinomas would be expected to be lower than that of normal endometrial tissue. However, in the present study, endometrioid carcinoma specimens did not differ significantly from normal endometrial tissue specimens in terms of E-cadherin expression. This may be due to the limited number of cases in both groups, the less invasive nature of endometrioid carcinomas, or the presence of expressed but dysfunctional E-cadherin in the endometrioid carcinoma group. In conclusion, the present study demonstrated that uterine non-endometrioid (papillary serous and clear cell) carcinomas were less likely to express E-cadherin than endometrioid carcinomas or normal endometrial tissue. This may help to explain the more aggressive behaviour of nonendometrioid carcinomas.
Conflicts of interest The authors had no conflicts of interest to declare in relation to this article.
• Received for publication 12 March 2008 • Accepted subject to revision 2 June 2008 • Revised accepted 6 October 2008 Copyright © 2009 Field House Publishing LLP References 1 Moreno-Bueno G, Hardisson D, Sarrió D, et al: Abnormalities of E- and P-cadherin and catenin
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9 Ascaño JJ, Frierson H Jr, Moskaluk CA, et al: Inactivation of the E-cadherin gene in sporadic diffuse-type gastric cancer. Mod Pathol 2001; 14: 942 – 949. 10 Motta FJ, Valera ET, Lucio-Eterovic AK, et al: Differential expression of E-cadherin gene in human neuroepithelial tumors. Genet Mol Res 2008; 7: 295 – 304. 11 Chetty R, Serra S, Asa SL: Loss of membrane localization and aberrant nuclear E-cadherin expression correlates with invasion in pancreatic endocrine tumors. Am J Surg Pathol 2008; 32: 413 – 419. 12 Shimoyama Y, Nagafuchi A, Fujita S, et al: Cadherin dysfunction in a human cancer cell line: possible involvement of loss of α-catenin expression in reduced cell–cell adhesiveness. Cancer Res 1992; 52: 5770 – 5774. 13 Goff BA, Kato D, Schmidt RA, et al: Uterine papillary serous carcinoma: patterns of metastatic spread. Gynecol Oncol 1994; 54: 264 – 268. 14 Hoffman K, Nekhlyudov L, Deligdisch L: Endometrial cancer in elderly women. Gynecol Oncol 1995; 58: 198 – 201. 15 Furuyama H, Arii S, Mori A, et al: Role of Ecadherin in peritoneal dissemination of the pancreatic cancer cell line, Panc-1, through regulation of cell to cell contact. Cancer Lett 2000; 157: 201 – 209. 16 Miyamoto S, Baba H, Kuroda S, et al: Changes in E-cadherin associated with cytoplasmic molecules in well and poorly differentiated endometrial cancer. Br J Cancer 2000; 83: 1168 – 1175.
Author’s address for correspondence Dr Tulin Yalta Pathology Department, Sivas City Hospital, Sivas, Turkey. E-mail:
[email protected]
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