Expression Of Cell-cell Adhesion Molecules Research Articles

P087 IFN-γ-Induced Intestinal Epithelial Cell Type-Specific Cytotoxic Responses of Human Enteroids: PANoptosis and The Protective Role of Selective JAK1 Inhibitors

Abstract Background Interferon-gamma (IFN-γ) plays an important role in the development of intestinal injury and inflammatory bowel disease (IBD). Using human intestinal organoid (enteorid) models, this study was investigated 1) what is the major response of intestinal epithelial cells (IECs) induced by IFN-γ to elucidate; 2) which programmed cell death (PCD) pathways cause IFN-γ-induced cell death on IECs in the absence of intestinal microbiota and immune cells; 3) how the expression of cell-cell adhesion molecules in epithelial cells is altered by IFN-γ; 4) how epithelial cell differentiation is altered by IFN-γ; 5) whether there is a difference in IFN-γ-induced PCD between different epithelial cell types; and 6) which molecules can block IFN-γ-induced PCD in IECs. Methods Using human enteroids, the major response of IECs induced by IFN-γ was evaluated, focusing on the IFN-γ-induced PCD pathway. The intestinal epithelial cell type-specific response to IFN-γ was assessed by bulk and single-cell RNA sequencing (RNA-seq). Furthermore, the molecules to block IFN-γ-induced PCD in IECs were evaluated. Results As the concentration of IFN-γ in the culture media increased, disruption of organoid structure, growth arrest, and cell death were observed in IFN-γ-treated enteroids on organoid reconstitution, MTT, and EdU assay. Bulk RNA-seq and western blot were identified the activation of pyroptosis, apoptosis, and necroptosis to form the collective inflammatory cell death pathway of PANoptosis. Single-cell RNA-seq indicated that IFN-γ altered epithelial cell differentiation in human enteroids, including expansion of the intstinal stem cell (ISC) population and depletion of the enterocyte population. PCD-associated gene expression was upregulated in enterocytes and goblet cells, but not in ISCs and paneth cells. Individual PCD inhibitors may be insufficient to prevent IFN-γ-induced cytotoxicity, whereas upadacitinib interferes with the downstream signaling of IFN-γ by inhibiting the activity of JAK1 kinase, resulting in effective blocking of PANoptosis. Conclusion PANoptosis is the major mechanism of IFN-γ-induced IEC damage, which was blocked by a selective JAK1 inhibitor. ECs were particularly susceptible to IFN-γ-induced PANoptosis. * This work was supported by the National Research Foundation of Korea(NRF) grant funded by the Korea government(MSIT) (2022R1A2B5B02002092 and 2021R1C1C2095192) and Future Medicine 2030 Project of the Samsung Medical Center (SMO1230011).

Time to go: neural crest cell epithelial-to-mesenchymal transition.

ABSTRACTNeural crest cells (NCCs) are a dynamic, multipotent, vertebrate-specific population of embryonic stem cells. These ectodermally-derived cells contribute to diverse tissue types in developing embryos including craniofacial bone and cartilage, the peripheral and enteric nervous systems and pigment cells, among a host of other cell types. Due to their contribution to a significant number of adult tissue types, the mechanisms that drive their formation, migration and differentiation are highly studied. NCCs have a unique ability to transition from tightly adherent epithelial cells to mesenchymal and migratory cells by altering their polarity, expression of cell-cell adhesion molecules and gaining invasive abilities. In this Review, we discuss classical and emerging factors driving NCC epithelial-to-mesenchymal transition and migration, highlighting the role of signaling and transcription factors, as well as novel modifying factors including chromatin remodelers, small RNAs and post-translational regulators, which control the availability and longevity of major NCC players.

Depot-medroxyprogesterone acetate reduces genital cell–cell adhesion molecule expression and increases genital herpes simplex virus type 2 infection susceptibility in a dose-dependent fashion

ObjectiveAnalyzing ectocervical biopsy tissue from women before and after they initiated use of depot-medroxyprogesterone acetate (DMPA), we previously reported this progestin reduces levels of the cell–cell adhesion molecule (CCAM) desmoglein-1 and increases genital mucosal permeability. We likewise saw treating mice with 1.0 mg of DMPA reduced vaginal CCAM expression and increased genital pathogen susceptibility. Herein, we used dose–response studies to delimit DMPA doses and serum MPA levels in mice associated with impaired genital mucosal barrier function and enhanced susceptibility to low-dose herpes simplex virus type 2 (HSV-2) infection. Study designWe compared genital CCAM expression, genital mucosal permeability, and susceptibility to genital inoculation with 103 plaque-forming units of HSV-2 among mice in estrus vs. after treatment with 0.01 mg, 0.1 mg, 0.3 mg, or 1.0 mg of DMPA. ResultsCompared to mice in estrus, DMPA treatment in a dose-dependent fashion significantly reduced desmoglein 1α (Dsg1a) and desmocollin-1 (Dsc1) gene expression, reduced DSG1 protein levels, and increased genital mucosal permeability to a low molecular weight molecule. While no mice infected with HSV-2 in estrus died, we respectively saw 50% and 100% mortality in mice administered 0.1 mg or 0.3 mg of DMPA. At time of infection, mean serum MPA levels in mice administered the 0.1 mg or 0.3 mg doses were 3.8 nM and 13.0 nM respectively (values comparable to trough and peak MPA serum levels in women using DMPA). ConclusionsMice with pharmacologically relevant serum MPA concentrations display significant changes in genital CCAM expression, genital mucosal barrier function, and HSV-2 susceptibility.

How do Single-Cell Properties Influence the Collective Mechanical Behavior of Confluent Tissues?

Biological tissues involved in important processes such as embryonic development, lung function, wound healing, and cancer progression have recently been shown, via a systematic analysis of tissue mechanics and cell displacement statistics, to be close to a disordered liquid-to-solid or “jamming” transition. We expect these jamming transitions to be very important for biological function. For example, due to collective effects, cell migration is greatly reduced in solid-like tissues and enhanced in fluid-like tissues. Therefore, we would like to know how cells might regulate properties such as expression of cell-cell adhesion molecules and myosin activity in order to change the macroscopic properties of the tissue as a whole. I will discuss a new theoretical framework that predicts how the jamming transition is governed by single-cell mechanics and persistent cell motility. I will discuss how our a priori theoretical predictions with no fit parameters are precisely realized in cell cultures from human patients with asthma, and discuss how these ideas might also be applied in heterogeneous cell populations to understand processes in embryonic development and cancer progression.

Salivary duct carcinoma with rhabdoid features: a salivary counterpart of pleomorphic lobular carcinoma of the breast.

To analyse the clinicopathological features and immunohistochemical characteristics of nine cases of salivary duct carcinoma (SDC) with rhabdoid features (SDCRF), representing a new, extremely rare type of salivary gland malignancy. We analysed 2511 cases of salivary gland tumour, clinicopathologically and immunohistochemically. The incidence of SDCRF was 0.4%. Eight patients were male. The age of patients ranged from 36 years to 85 years (mean, 61 years). SDC arose from the parotid glands and submandibular gland in six and three cases, respectively. Seven cases appeared as a carcinoma component of carcinoma ex pleomorphic adenoma cases. Six patients died of disease. Histologically, diffuse proliferations of non-coherent large ovoid or polygonal carcinoma cells with eosinophilic cytoplasm and eccentric nuclei were observed in all cases; such cytological characteristics were defined as 'rhabdoid features'. Immunohistochemically, all cases were positive for cytokeratin, gross cystic disease fluid protein-15, androgen receptor, and SMARCB1, seven cases were positive for HER2, and two cases were positive for epidermal growth factor receptor. However, all cases were negative for vimentin and myoepithelial markers. Eight cases showed no or aberrant expression of E-cadherin and β-catenin. The results suggest that SDCRF is an extremely rare subtype of SDC, and not a sarcomatoid variant of SDC. SDCRF is histologically unique, and is positive for SDC markers but negative for vimentin, unlike rhabdoid-type carcinomas arising from other organs. The morphogenesis of SDCRF is related to no or aberrant expression of cell-cell adhesion molecules. Therefore, SDCRF could be a salivary counterpart to pleomorphic lobular breast carcinoma.

Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype.

Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and β-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell-cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or pharmacological screening. Copyright © 2016 John Wiley & Sons, Ltd.

Suppression of lysyl-tRNA synthetase, KRS, causes incomplete epithelial-mesenchymal transition and ineffective cell‑extracellular matrix adhesion for migration.

The cell-adhesion properties of cancer cells can be targeted to block cancer metastasis. Although cytosolic lysyl-tRNA synthetase (KRS) functions in protein synthesis, KRS on the plasma membrane is involved in cancer metastasis. We hypothesized that KRS is involved in cell adhesion-related signal transduction for cellular migration. To test this hypothesis, colon cancer cells with modulated KRS protein levels were analyzed for cell-cell contact and cell-substrate adhesion properties and cellular behavior. Although KRS suppression decreased expression of cell-cell adhesion molecules, cells still formed colonies without being scattered, supporting an incomplete epithelial mesenchymal transition. Noteworthy, KRS-suppressed cells still exhibited focal adhesions on laminin, with Tyr397-phopshorylated focal adhesion kinase (FAK), but they lacked laminin-adhesion-mediated extracellular signal-regulated kinase (ERK) and paxillin activation. KRS, p67LR and integrin α6β1 were found to interact, presumably to activate ERK for paxillin expression and Tyr118 phosphorylation even without involvement of FAK, so that specific inhibition of ERK or KRS in parental HCT116 cells blocked cell-cell adhesion and cell-substrate properties for focal adhesion formation and signaling activity. Together, these results indicate that KRS can promote cell-cell and cell-ECM adhesion for migration.

Interplay between cadherins and α2β1 integrin differentially regulates melanoma cell invasion.

Background:Malignant transformation of melanocytes frequently coincides with an alteration in the expression of cell–cell adhesion molecules (cadherins) and cell-extracellular matrix proteins (integrins). How these two adhesion systems interplay to impact on cell invasion remains to be described in melanoma.Methods:Cell adhesion networks were localised by immunofluorescence in human primary cutaneous melanoma, metastatic melanoma in the lymph nodes, and melanoma cell lines. The role of these cell adhesion networks was assessed both in vivo, by analysing their impact on tumour growth in mice, and in vitro, with the use of functional tests including cell aggregation and cell migration.Results:We found that α2β1 integrin associates with both E-cadherin and N-cadherin to form two adhesive networks, distinguishable by the interaction—or not—of α2β1 integrin with type I collagen. N-cadherin/α2β1 integrin and E-cadherin/α2β1 integrin networks differently participated towards tumour growth in mice. The N-cadherin/α2β1 integrin network showed specific involvement in melanoma cell invasion and migration towards type I collagen. On the other hand, the E-cadherin/α2β1 network regulated cell–cell adhesion.Conclusions:This suggests that different signalling environments can be generated, depending on the type and/or local concentration of cadherin present in the adhesion complex, which potentially leads to differential cell responses. Further clarification of how these adhesive networks are regulated is fundamental to understanding important physiological and pathological processes such as morphogenesis, wound healing, tumour invasion and metastasis.

Bone Marrow Angiogenic Potential and Molecular Expression of Cell-Cell Adhesion Molecules and Matrix-Metallo Proteinases in Multiple Myeloma

Bone Marrow Angiogenic Potential and Molecular Expression of Cell-Cell Adhesion Molecules and Matrix-Metallo Proteinases in Multiple Myeloma

Abstract 179: Mechanical Assessment of the Drosophila Heart Reveals Conservation of Human Heart Failure Mechanisms

Rationale: Current models for investigating the mechanisms of heart failure are limited by low throughput or genetic intractability. Therefore, we have developed a nanoindentation method to measure the intrinsic stiffness of the intact, living Drosophila heart tube to assess mechanical function in this rapidly aging, genetically malleable system which has been shown to experience age-related decline in cardiac function. Objective: Fruit fly strains universally experience decline in fractional shortening with age due to decreased diastolic diameter, though severity varies between strains. We compared a strain with significant decline in diameter with age and one without to examine the hypothesis that diastolic decline is the consequence of increase passive tension due to myocardial stiffening with age. Quantitative PCR was employed targeted towards mechanosensitive molecules known to be overexpressed in failing human hearts. Methods and Results: Female yw flies experience ∼18% reduction in diastolic diameter with age (59.3 at 1 week to 48.4 µm at 5 weeks of age, p < 10-3) as calculated from m-mode kymographs of beating heart tube. Nanoindentation reveals 111% increase in stiffness (1.82 to 3.85 kPa, p < 10-10) proximal to the cardiac cell-cell junctions. In contrast, w1118 flies experience ∼6% decline in diastolic diameter (74.1 to 69.5 µm, p = 0.049) and do not stiffen with age (p = 0.1923). qPCR reveals increased expression of cell-cell adhesion molecules in yw, such as vinculin and cadherin. Expression levels remain steady or decline in w1118. Further analysis reveals that w1118 experiences separation between juxtaposed muscle layers (410 nm at 1wk, 448 nm at 5wks) indicating matrix adhesion defects not observed in yw (0 nm, p < 10-7 compared to 1wk w1118; 0 nm, p < 10-8 compared to 5wk w1118). Conclusion: These data indicate structural changes in the fly are a consequence of adherens-junction-mediated mechanical remodeling similar to that which is observed in the failing human heart. Matrix defects and the lack of remodeling in w1118 suggest a link between costamere and adherens signaling during aging. Cardiac matrix content and adhesion signatures likely direct cardiac remodeling and function.

Ex vivo bio-compatibility of honey-alginate fibrous matrix for HaCaT and 3T3 with prime molecular expressions

Honey's inherent compositional diversity, bio-compatibility and time tested therapeutic efficacy, especially in tissue repair as a topical agent, attract researchers towards harnessing its biomaterial potential particularly in developing matrix for tissue engineering applications. Hence, this study fabricates fibrous mat from optimum honey-alginate formulation and alginate solution using wet spinning technology. The physical and morphological properties of the scaffolds are assessed and finally their comparative biological performances are evaluated through in vitro studies on adherence, viability and prime molecular expression of HaCaT and 3T3 cells. The honey-alginate scaffold demonstrates better performance than that of alginate in terms of cellular adherence, viability and proper expression of cell-cell adhesion molecule (E-cadherin) and prime molecules of extra cellular matrix (Collagen I and III) by HaCaT and 3T3 respectively.

Regulation of spatiotemporal expression of cell–cell adhesion molecules during development of Dictyostelium discoideum

The social amoeba Dictyostelium discoideum is a simple but powerful model organism for the study of cell-cell adhesion molecules and their role in morphogenesis during development. Three adhesive systems have been characterized and studied in detail. The spatiotemporal expression of these adhesion proteins is stringently regulated, often coinciding with major shifts in the morphological complexity of development. At the onset of development, amoeboid cells express the Ca(2+) -dependent cell-cell adhesion molecule DdCAD-1, which initiates weak homophilic interactions between cells and assists in the recruitment of individuals into cell streams. DdCAD-1 is unique because it is synthesized as a soluble protein in the cytoplasm. It is targeted for presentation on the cell surface by an unconventional protein transport mechanism via the contractile vacuole. Concomitant with the aggregation stage is the expression of the contact sites A glycoprotein csA/gp80 and TgrC1, both of which mediate Ca(2+) /Mg(2+) -independent cell-cell adhesion. Whereas csA/gp80 is a homophilic binding protein, TgrC1 binds to a heterophilic receptor on the cell. During cell aggregation, csA/gp80 associates preferentially with lipid rafts, which facilitate the rapid assembly of adhesion complexes. TgrC1 is synthesized at low levels during aggregation and rapid accumulation occurs initially in the peripheral cells of loose mounds. The extracellular portion of TgrC1 is shed and becomes part of the extracellular matrix. Additionally, analyses of knockout mutants have revealed important biological roles played by these adhesion proteins, including size regulation, cell sorting and cell-type proportioning.

CDH3/P-cadherin overexpression in breast carcinomas: its regulatory mechanisms, the role in cell invasion, and the association with cancer stem cell properties

One of the most basic characteristics of cancer cells is the loss of cell-cell adhesion and acquirement of invasive properties. During cancer progression, the regulation and expression of cell-cell adhesion molecules (like cadherins) play a pivotal role. In most invasive tumors, E-cadherin is downregulated and N-cadherin is de novo expressed (known as epithelial-to-mesenchymal transition). However, in some invasive carcinomas, E-cadherin expression is maintained, with the concomitant de novo expression of another cadherin – P-cadherin. Their co-expression occurs frequently in breast carcinomas, which show a worst patient prognosis when compared with patients harboring tumors with loss of E-cadherin as single event. In the last ten years, our group has been mainly interested in understanding the role of P-cadherin overexpression in cancer cells, as well as the gene regulatory mechanisms behind its aberrant expression. P-cadherin has been extensively studied concerning its function and prognostic value in breast cancer. Its overexpression has been identified in 30% of invasive carcinomas, being highly associated with proliferative lesions of high histological grade and decreased patient survival. Recently, we showed that P-cadherin overexpression confers an invasive capacity to breast cancer cells, inducing the secretion of MMPs, which are responsible by the cleavage of its extracellular domain, giving rise to a P-cadherin soluble form. We proved that this fragment is a pro-invasive factor, which needs to be inhibited to render cancer cells non-invasive. Still, we identified the intracellular signaling pathway that regulates and activates the P-cadherin (CDH3) gene promoter, inducing P-cadherin overexpression in breast cancer cells. We found that an antiestrogen is able to increase CDH3 promoter activity, as well as to induce activating histone epigenetic modifications at putative C/EBPβ binding sites in the CDH3 gene promoter. We showed, for the first time, that C/EBPβ is able to regulate P-cadherin overexpression. Furthermore, our recent data supports the idea that P-cadherin has a role in cancer stem cell biology. We found that breast cancer cell lines, presenting the highest levels of P-cadherin, show the highest expression of the stem cell markers CD49f, CD44 and CD24, as well as ALDH1 activity. Importantly, we reproduced these results in primary tumors, where we found that P-cadherin overexpression occurs in breast carcinomas with a cancer stem cell phenotype (CD44+CD24-/low) and ALDH1 expression. These results are now being experimentally addressed by mammosphere, 2D and 3D culture in vitro assays, to be then confirmed in vivo. In conclusion, our results open new avenues in breast cancer treatment, since P-cadherin is likely to be a good therapeutic target for invasive carcinomas overexpressing this protein. Scientifically, our studies will help to understand better the pathogenesis of breast cancer and other cancer models involving cadherin’s alterations.

Oxygen deprivation inhibits basal keratinocyte proliferation in a model of human skin and induces regio‐specific changes in the distribution of epidermal adherens junction proteins, aquaporin‐3, and glycogen

It is generally accepted that hypoxia and recovery from oxygen deprivation contribute to the breakdown and ulceration of human skin. The effects of these stresses on proliferation, differentiation and expression of cell-cell adhesion molecules were investigated for the first time in an organotypic model of human skin. Fully stratified tissues were exposed to a time course of oxygen deprivation and subsequent reoxygenation. Regional changes in keratinocyte morphology, glycogen stores and cellular junctions were observed, with more differentiated layers of the epidermis exhibiting the first evidence of oxygen deprivation. Cellular swelling within the granular layer was concurrent with aquaporin-3 depletion. The keratinocyte adherens junction proteins E-cadherin and beta-catenin were dramatically decreased in a regio-specific manner throughout the epidermis following oxygen deprivation. In contrast, P-cadherin and the desmosomal proteins desmoplakin and desmoglein-1 were refractory to oxygen deprivation. Relative to normoxic controls, hypoxic tissues exhibited increased mRNA levels of the transcriptional repressor Slug; however, mRNA levels of the related transcriptional factor Snail were unaffected. All cellular and molecular changes were reversible upon reoxygenation. These results show that oxygen deprivation and reoxygenation exert differential effects on epidermal adhesion proteins and suggest a novel role for cadherins, beta-catenin, and Slug in hypoxia-induced junctional changes occurring in stratified squamous epithelium.

The Transcriptional Control of Trunk Neural Crest Induction, Survival, and Delamination

Trunk neural crest cells are generated at the border between the neural plate and nonneural ectoderm, where they initiate a distinct program of gene expression, undergo an epithelial-mesenchymal transition (EMT), and delaminate from the neuroepithelium. Here, we provide evidence that members of three families of transcription induce these properties in premigratory neural crest cells. Sox9 acts to provide the competence for neural crest cells to undergo an EMT and is required for trunk neural crest survival. In the absence of Sox9, cells apoptose prior to or shortly after delamination. Slug/Snail, in the presence of Sox9, is sufficient to induce an EMT in neural epithelial cells, while FoxD3 regulates the expression of cell-cell adhesion molecules required for neural crest migration. Together, the data suggest a model in which a combination of transcription factors regulates the acquisition of the diverse properties of neural crest cells.

CELL-CELL ADHESION MOLECULES AND SIGNALING INTERMEDIATES AND THEIR ROLE IN THE INVASIVE POTENTIAL OF PROSTATE CANCER CELLS

The highly variable natural history of prostate carcinoma may be reflected in heterogeneity of invasive potential between tumors. We have examined two prostate cancer cell lines of low invasive potential (CAHPV10 and PZHPV7) and three cell lines of high invasive potential (DU-145, PC-3, LNCapFGC), to determine whether specific adhesion molecule profiles correlated with their invasive behavior. Using an in vitro invasion assay, we demonstrated that DU-145, LNCapFGC and PC-3 cells were highly invasive compared with CA-HPV-10 and PZ-HPV-7 cells. LNCapFGC cells expressed high levels of E-cadherin, alpha-, beta- and gamma-catenin, desmoglein, desmoplakin and GSK3beta using immunoblotting. This was, in general, comparable to immunohistochemical staining. PC-3 cells had no E-cadherin or alpha-catenin, but expressed a high level of the HGF/SF receptor c-Met. In contrast, DU-145 cells were found to express E-cadherin and low levels for all other protein molecules, except c-Met. The DU-145 cell line also lacked alpha-catenin expression. In CA-HPV-10 and PZ-HPV-7 cells, there was no detection of APC, PECAM-1, P-cadherin or Wnt-1. DU-145, LNCapFGC and PC-3 cells formed cell-cell aggregates, which were reduced by inclusion of anti-E-cadherin antibody and the motogen HGF/SF. These results show that prostate cancer cells exhibit a diverse expression of cell-cell adhesion molecules and their signaling intermediates. The expression of these adhesion molecules bears an important relationship with the invasive phenotype of these cells.

Immunohistochemical studies of proliferative activity and expression of cell-cell adhesion molecules in ameloblastoma

Immunohistochemical studies of proliferative activity and expression of cell-cell adhesion molecules in ameloblastoma

E-cadherin and beta-catenin expression in breast medullary carcinomas.

The initial step of cancer invasion and metastasis is the escape of tumour cells from the primary site, involving disruption of normal cell-cell adhesion and E-cadherin (E-cad) and beta-catenin (beta-cat) down-regulation, as shown in various types of human malignancies including breast carcinomas. Medullary carcinomas are high grade and poorly differentiated tumours with syncytial typical pattern, and prognosis unexpectedly better than that in high grade breast carcinomas. In a series of 55 breast typical medullary carcinomas diagnosed according to the strict use of Ridolfi et al (Cancer 40: 1365-1385, 1977) criteria, E-cad and beta-cat were investigated using quantitative (SAMBA 2005 system) immunocytochemical assays on frozen sections. Results were compared to that obtained on paraffin sections and in a series (n=55) of grade 3 ductal carcinomas. It was shown that medullary carcinomas significantly (p<0.001) expressed more E-cad and beta-cat than grade 3 ductal carcinomas. E-cad and beta-cat correlated with high expression of P53, of c-erbB, and of Ki-67 antigens, and with lack of hormone receptors antigenic sites (p<0.001). It was concluded that favourable prognosis and syncytial pattern of typical breast medullary carcinomas likely results, at least partly, from a particular expression of cell-cell adhesion molecules, significantly limiting tumour growth and efficiently mastering the tumour cell dissemination, opposing to high proliferative activity (grade 3).

Gene regulation of cell adhesion molecules in neural morphogenesis.

A mounting body of evidence suggests that cell-cell adhesion molecules (CAMs) play critical roles in morphogenetic patterning and in laying down the initial tissue scaffold of the nervous system. Perturbations of CAM binding can lead to altered tissue pattern and interruption of tissue interactions to altered patterns of CAM expression. The combined factors that regulate the expression of CAMs and that drive early neural development are, however, largely unknown. We have hypothesized that the coordinate expression of homeobox (Hox) and paired box (Pax) transcription factors in various axes of the body plan leads to differential expression of particular CAM genes. Following this hypothesis, we have characterized the promoters and other regulatory regions of a number of genes specifying CAMs and have identified cis-regulatory elements that bind and respond to Hox and Pax proteins. Our recent experiments in vitro indicate, for example, that transcription factors encoded by Hox and Pax genes bind to specific DNA sequences in the N-CAM promoter and activate expression of the N-CAM gene. Experiments on transgenic mice carrying either the wild-type N-CAM promoter or a variant with mutations in the homeodomain binding sites (HBS) linked to a lac-Z reporter gene indicate that interactions with these elements are important in establishing and maintaining N-CAM expression in the spinal cord. We have also examined the regulatory sequences controlling expression of the gene for the neuron-glia adhesion molecule (Ng-CAM). Unlike N-CAM, which is also expressed in many non-neural sites, Ng-CAM is restricted to cells of the nervous system. After identifying this promoter for the Ng-CAM gene, we characterized a silencer region in the first intron of the gene that extinguishes the expression of Ng-CAM in fibroblasts but not in neuronal cells. Thus, a default mechanism can account for the restriction of Ng-CAM expression to the nervous system. The silencer region contains five neural-restrictive silencer elements and a binding site for the Pax3 protein, which also appears to have silencing activity. All of these findings suggest that Hox and Pax transcription factors can have both activating and silencing functions in regulating CAM gene expression. The general significance of these accumulated observations is that they connect the place-dependent expression of gene products capable of direct morphoregulation to the function of pattern-forming genes.

EXPRESSION OF E-CADHERIN, ALPHA-CATENINS AND BETA-CATENINS IN HUMAN GASTRIC CARCINOMAS - CORRELATION WITH HISTOLOGY AND TUMOR PROGRESSION

The expression of cell-cell adhesion molecule, E-cadherin and its associated proteins, alpha- and beta-catenins in human gastric carcinomas was examined by Western blotting. All the seven gastric carcinoma cell lines expressed E-cadherin except KATOIII, which was derived from pleural effusion of a scirrhous type stomach cancer or Borrmann's type-4 carcinoma. The expression of alpha-catenin was not detected in HSC43 derived from scirrhous carcinoma, while HSC39 expressed abnormal beta-catenin caused by genetic alteration. In gastric carcinoma cases, the levels of E-cadherin and alpha-catenin were significantly lower in poorly differentiated adenocarcinomas and scirrhous carcinomas when compared to other types of gastric carcinomas. Deeply invasive carcinomas expressed E-cadherin and alpha-catenin at lower levels. However, the expression level of alpha-catenin was not necessarily consistent with that of E-cadherin. One of 10 gastric carcinomas examined showed complete deletion of alpha-catenin gene in Southern blotting. beta-catenin was expressed at lower level in poorly differentiated adenocarcinomas than in well-differentiated adenocarcinomas. These findings suggest that reduction in the expression of E-cadherin and its associated molecules, catenins, is involved in the development and infiltrative growth of scirrhous type gastric carcinomas.