Role Of E-cadherin Research Articles

E-Cadherin and Cell Adhesion: a Role in Architecture and Function in the Pancreatic Islet

Background/Aims: The efficient secretion of insulin from β-cells requires extensive intra-islet communication. The cell surface adhesion protein epithelial (E)-cadherin (ECAD) establishes and maintains epithelial tissues such as the islets of Langerhans. In this study, the role of ECAD in regulating insulin secretion from pseudoislets was investigated. Methods: The effect of an immuno-neutralising ECAD on gross morphology, cytosolic calcium signalling, direct cell-to-cell communication and insulin secretion was assessed by fura-2 microfluorimetry, Lucifer Yellow dye injection and insulin ELISA in an insulin-secreting model system. Results: Antibody blockade of ECAD reduces glucose-evoked changes in [Ca²⁺]_i and insulin secretion. Neutralisation of ECAD causes a breakdown in the glucose-stimulated synchronicity of calcium oscillations between discrete regions within the pseudoislet, and the transfer of dye from an individual cell within a cell cluster is attenuated in the absence of ECAD ligation, demonstrating that gap junction communication is disrupted. The functional consequence of neutralising ECAD is a significant reduction in insulin secretion. Conclusion: Cell adhesion via ECAD has distinct roles in the regulation of intercellular communication between β-cells within islets, with potential repercussions for insulin secretion.

E-cadherin Interactions Regulate β-Cell Proliferation in Islet-like Structures

Islet function is dependent on cells within the islet interacting with each other. E-cadherin (ECAD) mediates Ca²⁺-dependent homophilic cell adhesion between b-cells within islets and has been identified as a tumour suppressor. We generated clones of the MIN6 β-cell line that stably over- (S) and under-express (αS) ECAD. Modified expression of ECAD was confirmed by quantitative RT-PCR, immunoblotting and immunocytochemistry. Preproinsulin mRNA, insulin content and basal rates of insulin secretion were higher in S cells compared to aS and control (V) cells. However, stimulated insulin secretory responses were unaffected by ECAD expression levels. ECAD expression did affect proliferation, with enhanced ECAD expression being associated with reduced proliferation and vice versa. Formation of islet-like structures was associated with a significant reduction in proliferation of V and S cells but not αS cells. These data suggest that ECAD expression levels do not modulate insulin secretory function but are consistent with a role for ECAD in the regulation of β-cell proliferation.

Depletion of E-Cadherin Disrupts Establishment but Not Maintenance of Cell Junctions in Madin-Darby Canine Kidney Epithelial Cells

E-cadherin forms calcium-dependent homophilic intercellular adhesions between epithelial cells. These contacts regulate multiple aspects of cell behavior, including the organization of intercellular tight junctions (TJs). To distinguish between the roles of E-cadherin in formation versus maintenance of junctions, Madin-Darby canine kidney (MDCK) cells were depleted of E-cadherin by RNA interference. Surprisingly, reducing E-cadherin expression had little effect on the protein levels or localization of adherens junction (AJ) or TJ markers. The cells underwent morphological changes, as the normally flat apical surface swelled into a dome. However, apical-basal polarity was not compromised, transmembrane resistance was normal, and zonula occludin protein 1 dynamics at the TJs were unchanged. Additionally, an E-cadherin/Cadherin-6 double knockdown also failed to disrupt established TJs, although beta-catenin was lost from the cell cortex. Nevertheless, cells depleted of E-cadherin failed to properly reestablish cell polarity after junction disassembly. Recovery of cell-cell adhesion, transepithelial resistance, and the localization of TJ and AJ markers were all delayed. In contrast, depletion of alpha-catenin caused long-term disruption of junctions. These results indicate that E-cadherin and Cadherin-6 function as a scaffold for the construction of polarized structures, and they become largely dispensable in mature junctions, whereas alpha-catenin is essential for the maintenance of functional junctions.

Dynamic analysis of hepatoma spheroid formation: roles of E-cadherin and β1-integrin

A spheroid is an in vitro multicellular aggregate that provides a microenvironment resembling that of normal tissue in vivo. Although cell adhesion molecules such as integrins and cadherins have been implicated in participating in the process of spheroid formation, little is known about the timing of their action. In this study, we have employed an image-based quantitative method to investigate the compactness of cell aggregates during hepatoma spheroid formation in a dynamic fashion. By modulating beta1-integrin and E-cadherin activity with specific blocking antibodies, ion chelators, and RGD-sequence-containing peptides, we show that these cell adhesion molecules mediate the formation of spheroids through the establishment of complex cell-cell and cell-extracellular matrix (ECM) interactions. The dynamics of spheroid formation can be separated into three stages. In the first stage, ECM fibers act as a long-chain linker for the attachment of dispersed single-cells to form loose aggregations through the binding of integrins. This is followed by a delay period in which cell aggregates pause in compaction, presumably because of the accumulation of sufficient amounts of E-cadherins. In the third stage, strong homophilic interaction of E-cadherins is a major factor for the morphological transition from loose cell aggregates to compact spheroids. These findings thus provide comprehensive information on the molecular mechanisms and dynamics of hepatoma spheroid formation.

P-142 Prognostic role of e-cadherin in resected non-small-cell lungcancer detected by immunohistochemistry on high-throughput tissue microarray

P-142 Prognostic role of e-cadherin in resected non-small-cell lungcancer detected by immunohistochemistry on high-throughput tissue microarray

Oncogenic Ras dominates overexpression of E-cadherin in malignant transformation of intestinal epithelial cells

BackgroundLoss of the adherens junction protein E-cadherin is a critical event during Ras-mediated transformation of intestinal epithelial cells. The purpose of our study was to determine if overexpression of E-cadherin prevents Ras-induced malignant transformation and suppresses cell growth. MethodsRat intestinal epithelial cells were constructed with a mutated human Ha-RasVal12 cDNA. In these cells, Ras is constitutively expressed or induced by addition of isopropyl-1-thio-B-D-galactopyranoside. Cells were transfected with a bicistronic retroviral system that expressed green fluorescent protein alone or this protein and human E-cadherin. E-cadherin expression was measured by Western blot analysis, and localization by immunofluorescence. Anchorage-independent growth in soft agar was examined as well as tumor growth in nude mice. ResultsAfter Ras induction, endogenous E-cadherin was downregulated, whereas overexpression of human E-cadherin was sustained. Oncogenic Ras dominated overexpression of E-cadherin by causing malignant transformation and E-cadherin mislocalization. Ras also promoted growth in soft agar and tumors in nude mice despite E-cadherin overexpression. ConclusionsOncogenic Ras subverts the tumor suppressor activity of E-cadherin in Ras-transformed intestinal epithelial cells by downregulating endogenous E-cadherin and mislocalizing transfected E-cadherin. The role of E-cadherin as a tumor suppressor in intestinal malignancies may be restricted by mutated or overactive Ras.

Signet ring carcinoma in Zollinger-Ellison Syndrom: the role of E-cadherin and gastrin in the pathogenesis of the disease

The early gastric cancer of the young with infiltrates of signet-ring cells, have been reported to be associated with germ-line mutations in the E-cadherin (CDH1) gene. It has also been demonstrated that cell-cell adherens junctions of the gastrointestinal tract mediated by the transmembrane protein E-cadherin, is regulated by gastrin and its prohormone progastrin. In this abstract we describe a patient with MEN1 with Zollinger-Ellison syndrome (ZES), who additionally developed a signet-ring carcinoma of the stomac, which is most-likely due to excessive local gastrin and progastrin levels within the gastric mucosa.

E-cadherin expression in primary carcinomas of the breast and its distant metastases.

Aberrant expression of E-cadherin has been associated with the development of metastases in patients with breast cancer. Even though the expression of E-cadherin has been studied in primary breast tumors, little is known about its expression at the distant metastatic sites. We investigate the relationship between E-cadherin expression in primary breast carcinoma and their distant, non-nodal metastases. Immunohistochemical analysis of E-cadherin was performed in tissues from 30 patients with primary invasive breast carcinoma and their distant metastases. E-cadherin expression was evaluated as normal or aberrant (decreased when compared with normal internal positive controls, or absent). Twenty-two (73%) invasive carcinomas were ductal, and eight (27%) were lobular. Of the primary invasive ductal carcinomas, 55% (12/22) had normal E-cadherin expression and 45% (10/22) had aberrant expression. All of the metastases expressed E-cadherin with the same intensity as (12 tumors) or with stronger intensity than (10 tumors) the corresponding primaries. Of the invasive lobular carcinomas, one of eight (12%) primary carcinomas and none of the metastases expressed E-cadherin in the cell membranes, but they accumulated the protein in the cytoplasm. Aberrant E-cadherin expression is frequent in invasive ductal carcinomas that progress to develop distant metastases. Distant metastases consistently express E-cadherin, often more strongly than the primary tumor. Invasive lobular carcinomas have a different pattern of E-cadherin expression, suggesting a different role for E-cadherin in this form of breast carcinoma.

Regulation of cell morphology and adhesion by the tuberous sclerosis complex (TSC1/2) gene products in human kidney epithelial cells through increased E-cadherin/beta-catenin activity.

We investigated the effects of overexpression of the tuberous sclerosis-1 and -2 (TSC1/2) gene products (hamartin and tuberin, respectively) in the human kidney epithelial cell line 293 with an inducible expression system. As we had observed previously in fibroblasts, 293 cells overexpressing hamartin and/or tuberin grew more slowly in vitro. However, here we also observed that the 293 overexpressing cells underwent a dramatic morphological change in which groups of cells formed compact clusters. The overexpressing cells also displayed decreased dissociation and increased reaggregation in vitro. These changes were found to be associated with an increased level of E-cadherin, which is known to regulate cell-cell interactions in epithelial cells, and of its binding partner beta-catenin. Consistent with the role of E-cadherin in these effects, we found that the observed changes in 293 cell morphology, dissociation, and adhesion were calcium-dependent, and were reproduced by overexpression of E-cadherin. In contrast, overexpression of TSC1 in rat embryo fibroblasts, which lack E-cadherin, failed to elicit the same changes as in 293 cells. We conclude that the hamartin/tuberin complex exerted a direct effect on the morphology and adhesive properties of 293 cells through regulation of the level and/or activity of cellular E-cadherin/beta-catenin.

Immunohistochemical study of the expression of E-cadherin in canine mammary tumours

To investigate the possible role of E-cadherin in canine mammary tumours 20 benign and 40 malignant tumours were evaluated by immunohistochemistry in formalin-fixed paraffin wax-embedded samples. In all the benign...

Mechanisms of monozygotic (MZ) twinning: a possible role for the cell adhesion molecule, E-cadherin.

Monozygotic (MZ) twins account for about one-third of all twins. Most MZ twins arise sporadically but there are familial occurrences. The causes of MZ twinning are largely speculative but involve both genetic and environmental factors. Genetic factors could include cell adhesion molecules and factors involved in growth and differentiation of the placenta and fetus. In this study, we examine the possible role of E-cadherin, a cell adhesion molecule, in the causation of twinning. A common polymorphism in the E-cadherin gene reduces expression of E-cadherin by up to 68%. We examined the E-cadherin polymorphism in DNA extracted from placental tissue from 121 MZ twins, 99 DZ twins, and 93 singletons. There was no significant difference in the distribution of the E-cadherin polymorphism in MZ twins although there was a slight over-representation of homozygotes for the polymorphism in MZ twins. We conclude that there is insufficient evidence to suggest that decreased E-cadherin expression alone is a causative factor in MZ twinning but it could be a contributory factor along with other genetic and environmental influences.

The role of E-cadherin in low-grade ductal breast tumourigenesis.

Grade I invasive ductal breast carcinomas have a specific pattern of genetic aberrations, namely gain of 1q and loss of 16q. This pattern is very similar to the changes seen in invasive lobular breast carcinomas (ILCs). The gene on 16q involved in ILC is known to be E-cadherin (CDH1). This study has investigated whether the same gene is responsible for grade I invasive ductal carcinoma (IDC), using allele imbalance analysis, mutation screening, and immunohistochemistry (IHC). The data suggest that despite the shared pattern of genetic aberrations seen in grade I IDC and ILC, CDH1 is not the target gene in low-grade ductal tumourigenesis.

Role of E-Cadherin, α-, β-, and γ-Catenins, and p120 (Cell Adhesion Molecules) in Prolactinoma Behavior

E-cadherin/catenin complex regulates cellular adhesion and motility and is believed to function as an invasion suppressor system. In a number of cancers, abnormal and reduced expression of E-cadherin/catenin complex is associated with tumor invasion and metastasis. Prolactinomas show frequent invasion on the surrounding structures, despite their histologically benign nature. Furthermore, gender-based differences in endocrine and surgical findings are found in patients with prolactinoma. To understand biological factors governing prolactinoma behavior, this study analyzed the expression of E-cadherin; α-, β-, and γ-catenins; p120; and cell proliferation marker MIB-1 labeling index in 13 invasive tumors (9 in men, 4 in women), 26 noninvasive tumors (4 in men, 22 in women), and 8 normal anterior pituitaries by immunohistochemistry. Immunostaining of E-cadherin; α-, β-, and γ-catenins; and p120 showed a membranous pattern of reactivity and generally stronger in normal pituitaries than in prolactinomas. Expression of E-cadherin and β-catenin was significantly lower in invasive than in noninvasive prolactinomas (P <.002 and P <.005, respectively), and reduced expression of E-cadherin and β-catenin was more frequent in invasive than in noninvasive prolactinomas (P <.001 and P <.05, respectively); in contrast, γ-catenin expression showed higher in invasive than in noninvasive prolactinomas (P <.05). Expression of E-cadherin was significantly lower in macroprolactinomas than in microprolactinomas (P <.01), and decreased expression of E-cadherin and β-catenin predicted high MIB-1 expression (P <.05). Moreover, the expression of E-cadherin and β-catenin was significantly lower in macroprolactinomas in men than in those in women (P <.01 and P <.02, respectively). No statistical correlations were observed between expression of α-catenin, p120, and clinicopathologic features. In conclusion, the reduction of E-cadherin and β-catenin expression was related to invasiveness and proliferative status of prolactinomas and correlated with the more aggressive behavior of prolactinomas in men compared with in women.

The Role of E-cadherin in the Motility and Invasion of Implantation Site Intermediate Trophoblast.

During early pregnancy, intermediate (extravillous) trophoblast infiltrates the basal plate and invades the spiral arteries, a physiological process required to establish the maternal-fetal circulation. Immunostaining studies have shown that differentiation of trophoblast into this invasive subpopulation is associated with down-regulation of E-cadherin expression. To study the function of E-cadherin in trophoblast in vitro, we restored E-cadherin expression in an E-cadherin negative human implantation site intermediate trophoblastic cell line, IST-1, using a recombinant adenovirus, E-cad/Ad5 which constitutively expresses E-cadherin. In contrast to the control IST-1 cells which were individual and pleomorphic in shape, E-cad/Ad5 transduced cells were cohesive, uniform, and round. The motility and invasiveness of E-cad/Ad5 transduced IST-1 cells, as compared with the control cells, was significantly reduced. These effects were contact-dependent and were attenuated by a function-perturbing anti-E-cadherin antibody. In conclusion, our results indicate that expression of E-cadherin in IST-1 cells results in a contact-mediated inhibition of motility and invasion and suggest an important role for E-cadherin down-regulation in the intermediate trophoblast during implantation.

The Role of E-cadherin in the Motility and Invasion of Implantation Site Intermediate Trophoblast

During early pregnancy, intermediate (extravillous) trophoblast infiltrates the basal plate and invades the spiral arteries, a physiological process required to establish the maternal–fetal circulation. Immunostaining studies have shown that differentiation of trophoblast into this invasive subpopulation is associated with down-regulation of E-cadherin expression. To study the function of E-cadherin in trophoblast in vitro, we restored E-cadherin expression in an E-cadherin negative human implantation site intermediate trophoblastic cell line, IST-1, using a recombinant adenovirus, E-cad/Ad5 which constitutively expresses E-cadherin. In contrast to the control IST-1 cells which were individual and pleomorphic in shape, E-cad/Ad5 transduced cells were cohesive, uniform, and round. The motility and invasiveness of E-cad/Ad5 transduced IST-1 cells, as compared with the control cells, was significantly reduced. These effects were contact-dependent and were attenuated by a function-perturbing anti-E-cadherin antibody. In conclusion, our results indicate that expression of E-cadherin in IST-1 cells results in a contact-mediated inhibition of motility and invasion and suggest an important role for E-cadherin down-regulation in the intermediate trophoblast during implantation.

T cell adhesion and cytolysis of pancreatic cancer cells: a role for E-cadherin in immunotherapy?

Pancreatic cancer is an aggressive and potent disease, which is largely resistant to conventional forms of treatment. However, the discovery of antigens associated with pancreatic cancer cells has recently suggested the possibility that immunotherapy might become a specific and effective therapeutic option. T cells within many epithelia, including those of the pancreas, are known to express the αEβ7-integrin adhesion molecule, CD103. The only characterised ligand for CD103 is E-cadherin, an epithelial adhesion molecule which exhibits reduced expression in pancreatic cancer. In our study, CD103 was found to be expressed only by activated T cells following exposure to tumour necrosis factor beta 1, a factor produced by many cancer cells. Significantly, the expression of this integrin was restricted mainly to class I major histocompatibility complex-restricted CD8+ T cells. The human pancreatic cancer cell line Panc-1 was transfected with human E-cadherin in order to generate E-cadherin negative (wild type) and positive (transfected) sub-lines. Using a sensitive flow cytometric adhesion assay it was found that the expression of both CD103 (on T cells) and E-cadherin (on cancer cells) was essential for efficient adhesion of activated T cells to pancreatic cancer cells. This adhesion process was inhibited by the addition of antibodies specific for CD103, thereby demonstrating the importance of the CD103→E-cadherin interaction for T-cell adhesion. Using a 51Cr-release cytotoxicity assay it was found that CD103 expressing T cells lysed E-cadherin expressing Panc-1 target cells following T cell receptor stimulation; addition of antibodies specific for CD103 significantly reduced this lysis. Furthermore, absence of either CD103 from the T cells or E-cadherin expression from the cancer cells resulted in a significant reduction in cancer cell lysis. Therefore, potentially antigenic pancreatic cancer cells could evade a local anti-cancer immune response in vivo as a consequence of their loss of E-cadherin expression; this phenotypic change may also favour metastasis by reducing homotypic adhesion between adjacent cancer cells. We conclude that effective immunotherapy is likely to require upregulation of E-cadherin expression by pancreatic cancer cells or the development of cytotoxic immune cells that are less dependent on this adhesion molecule for efficient effecter function.British Journal of Cancer (2002) 87, 1034–1041. doi:10.1038/sj.bjc.6600597 www.bjcancer.com© 2002 Cancer Research UK

E-Cadherin Expression Is Inversely Proportional to Tumor Size in Experimental Liver Metastases

Objective. The role of E-cadherin in the metastatic process remains controversial. In this study, we examined what role E-cadherin plays in relation to tumor size and microvessel density (MVD) of the liver metastatic lesion.Materials and methods. In F344 male rats, metastatic lesions were made by injecting RCN-9 cells into the spleen. We performed an immunohistochemical evaluation of E-cadherin and MVD in the metastatic lesions. According to a previous report, we focused on tumor sizes ranging from 2 to 3 mm, and the metastatic nodules were divided into two groups considering the value of the MVD.Results. The expression of E-cadherin was inversely proportional to tumor size (r = −0.882, P < 0.0001). MVD was directly proportional to metastatic tumor size (r = 0.726, P = 0.001). MVD in group A, in which the tumor size was 2.3 mm or more, was significantly higher than that in group B, in which the tumor size was less than 2.3 mm (3.13 ± 1.65% vs 1.75 ± 0.27%; P = 0.0069). The expression ratio of E-cadherin in group A was 28.21 ± 16.36%, while it was 82.33 ± 16.35% in group B (P = 0.0003).Conclusion. In liver metastasis, E-cadherin's function is preserved when the tumor is small and E-cadherin's expression is reduced in large tumors in which neovascularization is increased.

Clinical significance of serum E-cadherin levels in patients with haematological malignancies.

E-cadherin is a transmembrane glycoprotein that mediates Ca2+-dependent intracellular adhesion in normal epithelial cells. E-cadherin levels in serum are known to be significantly elevated in patients with epithelial carcinomas. However, the role of E-cadherin in haematopoietic cells is less clear. In this study, serum E-cadherin levels were therefore determined in patients with acute or chronic leukaemia, malignant lymphoma or myelodysplastic syndromes. Significant elevation of serum E-cadherin levels was detected in patients with haematological malignancies, and between types of acute leukaemias or subtypes of myelodysplastic syndromes, stages of malignant lymphoma, and phases of chronic leukaemia, respectively, compared with those in healthy adult volunteers. These findings suggest that E-cadherin might be expressed in malignant haematopoietic cells and might be useful as a diagnostic indicator in haematological malignancies.

E-cadherin is essential for gastric epithelial restitution in vitro: A study using the normal rat gastric mucosal cell line RGM1

The proliferation and migration of epithelial cells appear to have important roles in intercellular adhesion and the regeneration of gastric mucosal lesions. However, the role of E-cadherin, an important intercellular adhesion molecule, in restitution after gastric mucosal damage is unknown. This study was designed to investigate the possible role of E-cadherin in the regeneration of gastric mucosal lesions. Artificial small wounds were made in an RGM1 confluent monolayer sheet, and the healing process was monitored with or without the presence of different concentrations of anti-E-cadherin antibodies. E-cadherin mRNA and protein expression were determined by Northern blot analysis and immunostaining, respectively. Epithelial restitution in anti-E-cadherin antibody-treated monolayers was inhibited as compared with that in the controls without antibody. E-cadherin mRNA and protein expression were up-regulated transiently during the healing process. E-cadherin plays an important role during wound healing of gastric epithelial lesions and is crucial for sheet migration. (J Lab Clin Med 2001;138:236-42)

This month in J Lab Clin Med: Issue highlights for October 2001

This month in J Lab Clin Med: Issue highlights for October 2001