Fred T. Bosman

Reduced expression of the cadherin–catenin complex in oesophageal adenocarcinoma correlates with poor prognosis

The E‐cadherin–catenin complex is important for cell–cell adhesion of epithelial cells. Impairment of one or more components of this complex is associated with poor differentiation and increased invasiveness of carcinomas. Oesophageal adenocarcinomas causes early metastases, progress rapidly, and consequently have a poor prognosis. By means of immunohistochemistry, the expression of E‐cadherin and alpha‐ and beta‐catenin was studied in 65 oesophageal adenocarcinomas and 15 lymph node metastases. Expression of these proteins was evaluated with respect to clinico‐pathological parameters and patient survival. Expression of the proteins was strongly correlated. In carcinomas, reduced expression of E‐cadherin, alpha‐catenin, and beta‐catenin was found in 74, 60, and 72 per cent, respectively. Expression of E‐cadherin and alpha‐catenin correlated significantly with stage and grade of the carcinomas, whereas expression of beta‐catenin correlated only with grade. Reduced expression of all three proteins correlated with shorter patient survival. In contrast to grade, E‐cadherin and beta‐catenin were significant prognosticators for survival, independent of disease stage. We conclude that in oesophageal adenocarcinomas, decreased expression of E‐cadherin, alpha‐catenin and beta‐catenin are related events. Furthermore, expression of at least E‐cadherin and beta‐catenin is significantly correlated with poor prognosis.

Directional atherectomy for treatment of restenosis within coronary stents: clinical, angiographic and histologic results

OBJECTIVESnThe safety and long-term results of directional coronary atherectomy in stented coronary arteries were determined. In addition, tissue studies were performed to characterize the development of restenosis.nnnMETHODSnDirectional coronary atherectomy was performed in restenosed stents in nine patients (10 procedures) 82 to 1,179 days after stenting. The tissue was assessed for histologic features of restenosis, smooth muscle cell phenotype, markers of cell proliferation and cell density. A control (no stenting) group consisted of 13 patients treated with directional coronary atherectomy for restenosis 14 to 597 days after coronary angioplasty, directional coronary atherectomy or laser intervention.nnnRESULTSnDirectional coronary atherectomy procedures within the stent were technically successful with results similar to those of the initial stenting procedure (2.31 +/- 0.38 vs. 2.44 +/- 0.35 mm). Of five patients with angiographic follow-up, three had restenosis requiring reintervention (surgery in two and repeat atherectomy followed by laser angioplasty in one). Intimal hyperplasia was identified in 80% of specimens after stenting and in 77% after coronary angioplasty or atherectomy. In three patients with stenting, 70% to 76% of the intimal cells showed morphologic features of a contractile phenotype by electron microscopy 47 to 185 days after coronary intervention. Evidence of ongoing proliferation (proliferating cell nuclear antigen antibody studies) was absent in all specimens studied. Although wide individual variability was present in the maximal cell density of the intimal hyperplasia, there was a trend toward a reduction in cell density over time.nnnCONCLUSIONSnAlthough atherectomy is feasible for the treatment of restenosis in stented coronary arteries and initial results are excellent, recurrence of restenosis is common. Intimal hyperplasia is a nonspecific response to injury regardless of the device used and accounts for about 80% of cases of restenosis. Smooth muscle cell proliferation and phenotypic modulation toward a contractile phenotype are early events and largely completed by the time of clinical presentation of restenosis. Restenotic lesions may be predominantly cellular, matrix or a combination at a particular time after a coronary procedure.

EXPRESSION OF NUCLEOPHOSMIN/B23 IN NORMAL AND NEOPLASTIC COLORECTAL MUCOSA

Nucleophosmin/B23 is a 38u2009kD molecular phosphoprotein involved in ribosome assembly and transport. In view of the fact that nucleophosmin/B23 appears to be more abundant in tumour cells than in normal cells, the mRNA expression and immunohistochemical localization of nucleophosmin/B23 were investigated in 19 samples of non‐neoplastic mucosa, six adenomas, and 16 adenocarcinomas of the colorectum. Northern blot analysis revealed that nucleophosmin/B23 mRNA is expressed at a higher level in adenomas and carcinomas than in non‐neoplastic mucosa of the colorectum. Immunohistochemical staining of formalin‐fixed, paraffin‐embedded tissue sections after microwave antigen retrieval, using a nucleophosmin/B23‐specific monoclonal antibody, showed almost exclusively diffuse nuclear reactivity of a majority of the epithelial cells in non‐neoplastic mucosa: in adenomas, reactivity was almost exclusively nucleolar and in carcinomas, nuclear as well as nucleolar staining was observed. During mitosis, the immunoreactivity of nucleophosmin/B23 appears in the cytoplasm. The results indicate that the expression of nucleophosmin/B23 is higher in neoplastic than in non‐neoplastic colorectal mucosa. Furthermore, the pattern of nucleophosmin/B23 expression shifts from nuclear to nucleolar early in the adenoma–carcinoma sequence. The exact function of nucleophosmin/B23 in colorectal carcinogenesis remains to be determined.

Integrins: cell adhesives and modulators of cell function

SummaryIntegrins encompass a family of cell-surface molecules which play a crucial role in cell-cell and cell-extracellular matrix interaction. Of these heterodimeric transmembrane glycoproteins (consisting of an α and β chain) as yet at least 20 different types have been described, all with a different pattern of reactivity with extracellular matrix components. In this review the cell and tissue distribution of the integrins is discussed, with special emphasis on immunohistochemical localization of the β1 integrins and the α6β4 integrin. The β1 integrins comprise a subfamily in which eight α chains combine with one β (the β1) chain. The α2β1, α3β1 and α6β1 and the α6β4 integrins are expressed on a wide variety of epithelia on the basolateral surface or exclusively on the basal surface facing the basement membrane (e.g. α6β1 and α6β4). Leucocyte integrins, which share a common α2 chain, occur almost exclusively on white blood cells and their precursors. The vitronectin receptors, which share a common αv chain, occur in a wide variety of cell types. Integrins play a major role in the interaction of the cell with the extracellular matrix in order to create and maintain tissue architecture. It has become clear, however, that through integrin-ligand interaction cell function is also modulated. Furthermore, in pathological conditions integrins play a role of some significance. Integrins mediate leucocyte traffic in developing inflammatory processes and function in neoplastic growth when it comes to invasion and metastasis.

Apoptosis: Pathophysiology of Programmed Cell Death

In all normal tissues, cell proliferation and cell death are balanced. The physiology of normal cell death, which has become generally known as apoptosis or programmed cell death, has been intensely investigated in recent years. In this review the cell biology and biochemistry of apoptosis are discussed. Although apoptotic cells can be morphologically recognized, characteristic molecular features such as internucleosomal DNA fragmentation, and histochemical techniques such as in situ end labeling, facilitate the recognition of apoptosis. Many of the genes involved in the regulation of apoptosis, which include cell growth associated genes such as c-myc and p53, have been identified. It has become clear that the bcl-genes (more explicitly bcl-2 and bax) are important apoptosis regulators. The details of the mechanism of programmed cell death are, however, not completely unraveled. It has become clear that apoptosis plays an important role in organ and tissue development during embryogenesis. Examples are the morphogenesis of limbs from limb buds, the development of the central nervous system and the maturation of the hematopoietic and lymphatic systems. Hormonal regulation of cells and tissues is also partly executed through apoptosis. In a variety of disease apoptosis plays a role. In cancer, apoptosis is a crucial feature, and in the resolution of inflammatory reactions, apoptosis is essential. In neurodegenerative diseases, dysregulation of the cell death programme may play a role. Further elucidation of the role of apoptosis in these diseases may lead to new possibilities for treatment.

Immunohistochemical light and electron microscopy of basal laminar deposit

The formation of basal laminar deposit (BLD) is one of the histopathologic changes in the aging human macula. BLD is assumed to be an early stage of age-related macular degeneration. The location of BLD, between the RPE plasma membrane and its basement membrane and in the outer collagenous zone of Bruchs membrane, and its ultrastructure suggest that it is composed of excessive amounts of basement membrane material. The main components of basement membranes are type IV collagen, heparan sulfate proteoglycans (HSPG) and laminin. Labeled antibodies against these components can therefore be used for the identification and localization of basement membrane material by means of immunohistochemical techniques. In this study the presence of type IV collagen, laminin and HSPG was determined in aged human maculae by immunohistochemistry and immunoelectron microscopy. Tests for the presence of type VI collagen and fibronectin were also performed. We obtained 76 eyes from 68 human subjects at autopsy or after surgical enucleation for anteriorly located choroidal melanomas. The finely granular component of BLD stained positive with antibodies against type IV collagen, HSPG and laminin, but the long-spacing collagen component of BED did not. Neither component of BED was stained with antibodies against type VI collagen or fibronectin. We conclude that BLD consists partly of excess basement membrane material.

Prognostic value of p53, bcl-2, and c-erbB-2 protein expression in phaeochromocytomas.

Many studies have tried to discriminate malignant from benign phaeochromocytomas, but until now no widely accepted histological, immunohistochemical, or molecular methods have been available. In this study of 29 malignant and 85 benign phaeochromocytomas from 102 patients, immunohistochemistry was performed with antibodies to the tumour suppressor gene product p53 and the proto‐oncogene products bcl‐2 and c‐erbB‐2, using the avidin–biotin complex method. Malignant phaeochromocytomas showed a statistically significant higher frequency of p53 (p=0·042) and bcl‐2 (p=0·037) protein expression than their benign counterparts. The combination of both markers showed an even higher significance (p=0·004), to which both markers contributed equally. Overexpression of c‐erbB‐2 was associated with the occurrence of familial phaeochromocytomas (p=0·001), but no difference was found between benign and malignant cases. In conclusion, p53, bcl‐2, and c‐erbB‐2 all appear to be involved in the pathogenesis of a proportion of phaeochromocytomas. Immunoreactivity to p53 and bcl‐2 proteins may help to predict the clinical behaviour of phaeochromocytomas. Copyright

Expression of Bcl-2 protein in hyperplastic polyps, adenomas, and carcinomas of the colon.

The proto‐oncogene Bcl‐2 encodes a protein that protects cells from programmed cell death (apoptosis). The protein is expressed in the proliferative compartment of several normal tissues, including normal colonic crypts. The aim of this study was to test Bcl‐2 expression in colorectal neoplasms, assuming that, as a regulator of apoptosis, it might be involved in the progression from adenoma to carcinoma. To this end, Bcl‐2 reactivity was tested by immunohistochemistry in hyperplastic polyps, colonic adenomas, and carcinomas and its expression was compared with staining for the proliferation‐associated Ki‐67 antigen, using the MIB‐1 antibody. Bcl‐2 expression occurred in 2 out of 10 hyperplastic polyps and in 31 out of 35 (tubular, villous, and tubulovillous) adenomas, irrespective of their degree of dysplasia. Of ten carcinomas, only three were focally Bcl‐2‐positive, all moderately to well differentiated. In two of four carcinomas in Bcl‐2‐positive adenomas, no Bcl‐2 staining was observed. High numbers of MIB‐1‐positive cells were found in all hyperplastic and neoplastic lesions, without apparent correlation between proliferation and Bcl‐2 expression. These findings suggest that in the pathogenesis of hyperplastic polyps, increased crypt cell proliferation is primarily involved, but in some lesions decreased apoptosis may play a role. Furthermore, the increased Bcl‐2 expression in adenomas but not in the majority of the carcinomas suggests either that decreased apoptosis is not usually involved in the pathogenesis of these lesions or that the regulation of apoptosis in colorectal epithelia involves additional regulatory factors.

In vivo and in vitro invasion in relation to phenotypic characteristics of human colorectal carcinoma cells

In this study we investigated the tumorigenicity, growth pattern and spontaneous metastatic ability of a series of nine human colorectal carcinoma cell lines after subcutaneous and intracaecal xenografting in nude mice. CaCo2 cells were found to be poorly tumorigenic to non-tumorigenic in either site; the other cell lines were tumorigenic in both sites. SW1116, SW480 and SW620 did not show local invasive in the NCI-H716 and LS174T cells were both invasive in the caecum, but only NCI-H716 was invasive in the subcutis. HT29 and 5583 (S and E) cells were invasive in the caecum and from that site metastatic to the lungs and/or the liver. HT29 and 5583S cells were both invasive in the subcutis, but 5583E cells were not. Of each category of in vivo behaviour in the caecum, one cell line was further investigated with regard to invasion in vitro (into embryonic chick heart fragments), E-cadherin expression in vivo and in vitro and in vitro production of u-PA and t-PA. These parameters were chosen in view of their purported role in extracellular matrix degradation and intercellular adhesion, which are all involved in the invasive and metastatic cascade. Invasion in vitro was not predictive for invasion or metastasis in vivo. In the cell line which showed invasion in embryonic chick heart tissue, heterogeneous E-cadherin expression was observed in vitro together with a relatively high production of u-PA. The non-invasive cell lines showed in vitro homogeneous expression of E-cadherin with a relatively low production of u-PA. In vivo expression of E-cadherin was either absent or heterogeneous. We conclude that: (1) colorectal carcinoma xenografts show site-specific modification of in vivo invasive and metastatic behaviour; (2) invasion in vitro does not correlate with invasion and metastasis in vivo; (3) in vitro non-invasion might be associated with homogeneous E-cadherin expression and low production of u-PA; (4) E-cadherin expression in vitro differs from E-cadherin expression in vivo. The results support the notion that the microenvironment in which cancer cells grow is one of the factors involved in the regulation of invasive and metastatic behaviour.

Basement membrane heterogeneity

Cette etude resume les caracteristiques communes a toutes les membranes basales et discute certains aspects de leur heterogeneite