Immunohistochemical analysis on normal nephrogenesis and Wilms' tumour using monoclonal antibodies reactive with lymphohaemopoietic antigens

Summary Adult and fetal human kidneys were investigated for the reactvities of monoclonal antibodies, BA-1, BA-2 and BA-3 against human leukocytes. In developing metanephros, their reactivities changed reflecting the stage of nephrogenesis. Thus BA-1 stained both metanephric blastema and ureteric bud. Glomerular and proximal tubular development was characterized by the disappearance of BA-1 reactive antigen and the appearance of CALLA defined by BA-3. Immuno-electron-microscopically CALLA was solely located on the glomerular epithelial membrane and on the microvilli of the proximal tubules. BA-2 constantly stained ureteric bud-derived tissues. These observations were applied for the analysis of histogenesis of Wilms' tumour. Tumour blastema of the classical type reacted only with BA-1. Epithelial components in the classical as well as the epithelial type reacted both with BA-I and BA-2. CALLA was only detectable in glomeruloid and the connecting tubular structures, while these were unstained by BA-1. Stromal components in the classical and the sarcomatous type did not express any of these antigens. Staining patterns were identical in surgically removed and xenotransplanted tumours. These studies establish that each component of Wilms' tumour can clearly be interpreted regarding its histogenesis and that epithelial components corresponding to proximal tubules are identified in Wilms' tumour.     

Neuroglial tissue in partially cystic Wilms'' tumour

Wilms' tumour (nephroblastoma), the commonest malignant renal tumour of childhood, is thought to be derived from metanephric blastema and a majority of cases show the classical triphasic pattern of blastema, stroma and epithelium. There is usually little difficulty in making a correct histological diagnosis, but problems can arise due to the extraordinary potential of metanephric blastema to differentiate into a wide variety of heterologous tissues. Here we report, probably for the first time, two cases of Wilms' tumour which contained prominent quantities of neuroglial tissue. The classification of these cases as Wilms' tumours rather than renal teratomas is discussed.     

An analysis of histology and DNA-ploidy in primary wilms tumors and their metastases and a study of the morphological effects of therapy

Summary In children with Wilms' tumours the length of survival is greatly influenced by success in preventing or controlling metastatic disease. The current study focuses on the morphological aspects of metastases when compared with the primary tumour. In 8 patients it appeared that blastema is the most likely component to metastasize, whereas epithelial and stromal components were hardly, if at all, represented in metastases. Furthermore, flow cytometric DNA ploidy determinations on 4 cases showed that both the primary tumours and the metastases had stemlines in the diploid and low aneuploid (hyperdiploid) range. Finally, in four cases the influence of therapy on morphology of the primary tumours was analyzed. In these cases blastema seemed to be the component most sensitive to therapy. Thus, blastema seems to play a central role in prognosis of Wilms' tumours; either reacting to therapy or, if insensitive, by metastasizing.This study was supported by the Groningen Foundation of Pediatric Oncology and partly by grant GUKC 84-6 of the Netherlands Cancer Foundation (Koningin Wilhelmina Fonds).     

Human lung tumours: a correlation of antigenic profile with histological type

Fifty-four human lung tumours have been immunostained with a large panel of monoclonal antibodies including reagents against cytokeratins, prekeratins, epithelial membrane antigen (EMA), carcinoembryonic antigen (CEA) and neural antigens. These results have been compared with the histological types of tumour using the current WHO classification scheme. The most striking finding of this study was the considerable overlap of antigenic profile between different histological types of tumour. This suggests that there may be a greater underlying similarity between different histological categories of lung tumour than has hitherto been assumed. Secondly it was evident that immunostaining highlighted areas of different morphology within many tumours emphasizing the heterogeneous differentiation patterns seen in lung cancers. The present study supports the viewpoint that lung tumours arise from a common stem cell and that these neoplasms represent a single tumour with a tendency to differentiate along one or more pathways.     

Study of childhood renal tumours using antisera to fibronectin, laminin, and epithelial membrane antigen.

Using a peroxidase-antiperoxidase staining procedure, formalin fixed paraffin embedded sections of fetal and normal kidney; benign (mesoblastic nephroma); and malignant tumours (Wilms' tumour, clear cell renal carcinoma, rhabdoid renal tumour, and bone metastasising renal tumour of childhood (BMRTC] were examined for their reactivity with antisera to fibronectin, laminin, and epithelial membrane antigen. Mesoblastic nephroma contained fibronectin but no laminin. Most Wilms' tumours lacked both fibronectin and laminin; 50% of rhabdoid renal tumours were positive for fibronectin and laminin--rhabdoid tumours as recognised morphologically may, in fact, be two separate entities. BMRTC and clear cell renal carcinoma lacked both fibronectin and laminin. Epithelial membrane antigen was present in most of the tubular Wilms' tumour but absent in blastemal Wilms' tumours. The presence of epithelial membrane antigen in rhabdoid tumours was surprising, as histologically, this type of tumour shows no sign of epithelial differentiation. Epithelial membrane antigen antiserum stained clear cell renal carcinomas: epithelial membrane antigen is found in the distal and not the proximal tubules of fetal and normal kidneys. Thus an obvious interpretation is that clear cell renal carcinomas originate from distal rather than from proximal tubules, as has always been thought. On the basis of these results and data from other published findings some possible histogenetic origins of childhood renal tumours were proposed.     

A monoclonal antibody stains blastemal but not tubular components of Wilms' tumour

The monoclonal antibody PAL-E is specific for endothelial cells in a wide variety of normal and tumour tissue. In normal kidney, PAL-E reacts exclusively with the endothelium of non-glomerular blood vessels. In Wilms' tumour, binding of PAL-E was not restricted to the endothelium; staining of blastemal cells was observed in seven out of eight cases examined. Mesenchymal and tubular components, if present in Wilms' tumour, were negative. In contrast, a monoclonal antibody to Factor VIII-related antigen (RFF-8-R-1) bound only to endothelial cells in these tumours. In fetal kidney, PAL-E binding showed a wider distribution than in adult kidney and both stromal and glomerular capillaries were stained. Tubules and non-endothelial stromal cells were negative. These results indicate that the reactivity of the monoclonal antibody, PAL-E, is not restricted to cells of endothelial origin in all tissues. The implications of these findings for the differentiation of Wilms' tumour are discussed.     

Immunocytochemistry of malignant mesothelioma: OV632 as a marker of malignant mesothelioma.

In pleural or ascitic effusions the cytomorphological distinction of adenocarcinoma cells, reactive mesothelial cells, and malignant mesothelioma cells often causes a diagnostic dilemma. The value of immunocytochemistry was investigated on cytological smears of 24 well-established cases of malignant mesothelioma, a selected series of 31 metastatic adenocarcinomas, and 20 smears of patients without known malignancy. In these smears we scored the immunoreactivity with a panel of four monoclonal antibodies. In addition to antibodies for epithelial membrane antigen (EMA) and carcinoembryonic antigen (CEA), the monoclonal antibody MOC31 and the ovarian carcinoma specific antibody OV632 were incorporated in the panel. With none of these four antibodies was immunostaining of reactive mesothelial cells found. CEA- and MOC31-positive tumour cells were frequent in metastatic adenocarcinomas, but occurred rarely in malignant mesotheliomas. EMA-positive tumour cells were found in all metastatic adenocarcinomas (100 per cent) and in most malignant mesotheliomas (83 per cent). In addition to the expected reactivity of OV632 with ovarian carcinomas, 22 of 24 malignant mesotheliomas contained immunopositive tumour cells, while only a small proportion of non-ovarian adenocarcinomas reacted with this antibody. This selective staining of malignant mesothelioma cells, but not reactive mesothelial cells, with OV632 now permits the positive identification of malignant mesothelioma cells in male patients.     

Choroid plexus papillomas: an immunohistological study of 16 cases

Eleven benign and five malignant choroid plexus papillomas in children and adults were studied immunohistologically with a panel of antibodies against glial fibrillary acidic protein, S-100 protein, vimentin, desmin, epithelial membrane antigen and two different cytokeratins (LP34 and CAM 5.2). Glial fibrillary acidic protein was focally present in epithelial tumour cells, in cells within solid areas and in clusters of cells within the stroma. S-100 protein was diffusely present in tumour cells with focal accentuation. Vimentin was present in all cases, the epithelial tumour cells demonstrating strong and diffuse positivity with perinuclear accentuation; malignant tumours, however, showed stronger positivity than benign ones. Desmin was negative in all tumours. Epithelial membrane antigen and cytokeratin (LP34) were demonstrated in four of five malignant tumours but were absent in the benign ones; CAM 5.2 reacted with four of five malignant tumours and also reacted with eight of the 11 benign ones. The significance of these findings is discussed in respect of the ontogeny of these tumours.     

Diagnosing tumours on routine surgical sections by immunohistochemistry: use of cytokeratin, common leucocyte, and other markers

Tumours of uncertain tissue of origin were investigated by immunohistochemistry on formalin fixed paraffin embedded sections. Two antibodies--PD7/26, an anti common leucocyte antigen, and CAM5.2, an anticytokeratin--recognised most lymphomas and carcinomas, respectively: 88% of these tumours were identified by the two antibodies alone. These antibodies permitted the separation of the cases into groups: positive with CAM5.2, positive with PD7/26, and a third comprising those negative with both. The negative group contained other tumours and a small number of carcinomas and lymphomas; many of the lymphomas were, apparently, of histiocytic origin. Comparison of CAM5.2 with other epithelial markers showed that it was the most effective. Some further classification of the tumours was carried out with a panel of organ and cell specific antibodies: mesotheliomas were recognised by their pattern of reactivity with epithelial markers. Overall, the tumour type was determined in 90% of cases. Immunohistochemistry performed as described can be a potent aid to the diagnostic histopathology of tumours.     

The in vitro growth, heterotransplantation, and immunohistochemical characterization of the blastemal component of Wilms' tumor

Wilms' tumor has been proposed to originate from a developmental abnormality of the metanephric blastema. This undifferentiated component of Wilms' tumors has previously eluded efforts for in vitro growth. Blastema from a "classical" Wilms' tumor was transplanted into nude mice and passaged through 12 generations of heterotransplantation. Tumors from heterotransplants were grown for 12 serial passages in a serum-free growth medium supplemented with hormones and conditioned media from human kidney proximal tubule cells. The blastema initially grew on a collagen-fetal calf serum matrix as multicellular spheroids, and the cells proliferating from the rim of the spheroids had a flattened shape. Pulse-labeling with bromodeoxyuridine (BrdU) identified the proliferating cell population as blastemal in origin. Except for a loss of extracellular matrix, ultrastructural studies demonstrated morphologic similarities in the cultured cells, compared with the primary tumor and heterotransplants. Lectin histochemical stains for the peanut lectin (PNA) and immunohistochemical stains for cytokeratin (CYTO), vimentin (VIM), and epithelial membrane antigen (EMA) were performed on the original tumor, successive heterotransplants, and cells grown in vitro. The PNA stained the surface of the blastemal cells after sialidase digestion in the original tumor, heterotransplants, and cultured cells. The blastema of the original tumors was negative for CYTO and EMA but reactive for vimentin. This lack of differentiation was maintained in heterotransplants through 12 passages. However, blastemal cells demonstrated coexpression of CYTO and VIM intermediate filaments when grown in a serum-free medium on a matrix material. These studies demonstrate that the blastemal component of Wilms' tumor can be successfully grown in culture, passaged in nude mouse heterotransplants, and shown to undergo early stages of blastemal differentiation in vitro by growth in serum-free medium. This in vitro system provides a model for testing the factors that influence the growth and differentiation of the blastemal component of Wilms' tumors.     

Immunoreactive cytokeratins in plasmacytomas

Fourteen plasma cell tumours, including examples of solitary plasmacytoma and multiple myeloma, were studied with a panel of antibodies reactive in formalin-fixed, paraffin wax-embedded tissue. Each case showed immunoglobulin light chain restriction. Five tumours were reactive with antibodies to cytokeratin. Of these five cases, four were negative with antibodies to leucocyte common antigen and only one was weakly positive. Anti-cytokeratin reactivity by plasma cell tumours is more common than was originally anticipated and represents an important diagnostic pitfall.     

Wilms' tumour in adults: a report of three cases

Wilms' tumour is the most common pediatric renal neoplasm. Its occurrence in adults is rare, less than 1% of all nephroblastomas have been reported after the age of 15 years of age. We report 3 cases of Wilms' tumour in adults, their clinico-pathological profile and review the literature including the cases reported from India. Patients were young adults, their age ranging from 25 to 32 years. The largest tumour dimension was between 11 cm to 17 cm. Two cases were blastema predominant while the third case had typical triphasic histology. All cases were stage II (National Wilms' Tumour Study), favourable histology. Although Wilms' tumour in adults is morphologically similar to its childhood counterpart, it often responds poorly to the combination chemotherapy. The criteria for its diagnosis are strict and should be adhered to for its correct identification.     

Pulmonary blastoma. An immunohistochemical analysis with comparison with fetal lung in its pseudoglandular stage

Pulmonary blastomas are believed to be mixed epithelial and mesenchymal tumors that recapitulate the developing lung at 10-16 weeks gestation. The authors compared nine blastomas with ten fetal lungs in the pseudoglandular stage of development with a panel of antibodies to various lung antigens to evaluate immunophenotypic homology. Both blastomas and embryonal lungs showed expression of cytokeratin, epithelial membrane antigen, and carcinoembryonic antigen in their epithelial elements, and both contained scattered chromogranin-positive neuroendocrine cells. Rare surfactant-producing and Clara cell antigen-elaborating cells were identified in both groups. The mesenchymal components of blastomas and fetal lung showed smooth muscle, myofibroblastic, and blastematous differentiation. The blastematous elements demonstrated vimentin and keratin coexpression in four cases, providing some support for the contention that pluripotential blastema may give rise to the epithelial and mesenchymal elements of the distal lobule.     

Pigmented neuroectodermal tumour of infancy: an immunohistochemical study

The pigmented neuroectodermal tumour of infancy is a rare neoplasm of uncertain histogenesis which, in the majority of cases, arises in the maxilla and pursues a benign course. Currently, it would be classified in the group of peripheral primitive neuroectodermal tumours. Histologically it is composed of two principal cell types: neuroblast-like and melanocyte-like. Three typical cases are presented herein, which appear to be the first examined with a panel of antibodies. The neuroblast-like cells labelled positively for neurone-specific enolase but were negative for S-100, neurofilaments, glial fibrillary acidic protein, vimentin, cytokeratin, epithelial membrane antigen (EMA) and carcinoembryonic antigen (CEA). The melanocyte-like cells stained positively for neurone-specific enolase, vimentin and cytokeratin but were negative for S-100, neurofilaments, glial fibrillary acidic protein, EMA and CEA. The significance of these findings is discussed in the light of previous suggestions about the differentiation that these tumours show.     

Primary extrarenal Wilms'' tumour: identification of a putative precursor lesion

We report a case of primary extrarenal Wilms' tumour which, on histological examination, revealed a zone of hyalinized blastema adjacent to, and within the tumour capsule. The tumour showed a predominant stromal component. The presence of the hyalinized blastema adjacent to the tumour raises the possibility that some cases of extrarenal Wilms' tumour may have a precursor lesion.     

Malignant renal rhabdoid tumour. Immunohistochemical and ultrastructural studies

In an effort to establish their possible histogenesis, three cases of renal rhabdoid tumour and their metastases were studied both by a battery of polyclonal and monoclonal antibodies using the avidin-biotin-peroxidase complex technique and by electron microscopy. Vimentin was demonstrated in renal rhabdoid tumour in two cases and in all metastatic deposits. Cytokeratin (39, 43 and 50 kD) was not demonstrable in the three renal rhabdoid tumours, but was strongly positive in all metastatic lesions in one case. Epithelial membrane antigen was present in one renal rhabdoid tumour and in pulmonary metastases in two cases. Ultrastructural study showed epithelial differentiation in all tumours: basal lamina and convergent tight junctions were demonstrated; intracytoplasmic intermediate filaments were present in all primary and metastatic tumours. Rhabdoid tumours thus exhibited heterogeneous immunophenotypic expression suggesting that they are derived from mesenchymal cells which are capable of differentiating into epithelial cells.     

Immunohistochemistry of ovarian granulosa cell tumours

Summary Paraffin-embedded material of 47 ovarian tumours primarily diagnosed as granulosa cell tumours, including 2 cases of juvenile granulosa cell tumour, were studied immunohistochemically for the presence of intermediate filament proteins, epithelial membrane antigen and tumour markers. Forty-one lesions, including the 2 juvenile granulosa cell tumours, were vimentin positive, while keratin and epithelial membrane antigen expression could not be detected. Six tumours primarily diagnosed as poorly differentiated malignant granulosa cell tumours were vimentin negative, showed a mild to moderate positivity for keratin and intense positivity with the anti-epithelial membrane antigen antibody. These latter tumours were therefore classified as undifferentiated ovarian carcinomas, corresponding to their significantly poorer prognosis and shorter survival when compared with the granulosa cell tumours. Two of these six tumours were positive for carcino-embryonic antigen. Two small cell carcinomas of the ovary studied in addition expressed keratin in a proportion of tumour cells while no epithelial membrane antigen or vimentin was detectable. None of the tumours tested for alpha-fetoprotein, human chorionic gonadotrophin, human placental alkaline phosphatase and human placental lactogen, were positive.The data indicate the value of antibodies directed against intermediate filament proteins and epithelial membrane antigen to distinguish granulosa cell tumours from poorly differentiated carcinomas, a worthwhile distinction considering the much better prognosis of granulosa cell tumours.     

Expression and localization of HGF and met in Wilms' tumours.

A number of growth factors and cognate receptors that contribute to normal kidney development have been shown to play roles in the pathogenesis of Wilms' tumours. Expression of both hepatocyte growth factor (HGF) and its tyrosine kinase receptor met has been demonstrated in normal tissues and their neoplastic counterparts, implicating these factors in normal development and tumour progression. HGF and met expression has not been studied in Wilms' tumour. Since HGF and met function in a paracrine fashion by regulating tubulogenesis in normal kidney development, they could be involved in the pathogenesis of Wilms' tumour, in which tubular formation is dysplastic. In the present study, a series of ten homotypic (consisting of blastemal, epithelial, and stromal elements) and ten heterotypic (consisting of triphasic histology and a muscle component) Wilms' tumour cases were examined for expression of HGF and met, using in situ hybridization, immunohistochemistry, and western blot analysis. Relatively high met message and protein expression, compared with normal kidney, were evident in homotypic and heterotypic tumour blastemal, epithelial, and rhabdomyoblastic cells and a 145 kD met polypeptide was found in all tumours, with a few cases also expressing the 170 kD precursor form. No apparent alterations of the met receptor were observed. Similarly, HGF protein was also abundantly expressed in blastemal, epithelial, and rhabdomyoblastic cells of the homotypic and heterotypic Wilms' tumours and a 69 kD HGF polypeptide was demonstrated by western blot analysis. Immunohistochemistry for the Ki-67 proliferation marker indicated that the pattern of Ki-67 expression correlated with the HGF and met pattern of expression in both homotypic and heterotypic tumours. These results reveal, for the first time, significant co-expression of met/HGF in Wilms' tumours, with a correspondingly high proliferative index in the same cell groups.     

Malignant triton tumor (immunomorphological research)

Immunohistochemical study with the use of PAP method on paraffine sections of two cases of triton tumour one of which developed against the background of Recklinghausen disease. The presence of vimentin and S-100 protein in the neurofibromatous cells and spindle-cell component of malignant triton tumour is shown. The muscle differentiation markers (desmin and myoglobin) are found in the polymorph-cell component. The positive reaction of the cytoplasmic processes of some cells in the neurofibromatous tissue to the antibodies against GFAP (glial fibrillar acid protein) is discussed. It is suggested that the number of S-100 protein-positive and vimentine-positive cells in the peripheral nerve sheath tumours reflects the degree of their differentiation. Hypotheses of the histogenesis of these rare tumours are discussed. The panel of antibodies for the differential diagnosis of malignant triton tumour with rhabdomyosarcoma, malignant schwannoma and other soft tissue tumours is proposed.     

Tumour marker immunoreactivity in adenocarcinoma.

To determine whether variations in the expression of tumour related antigen can predict the origin of tumours, the immunoreactivity of a series of adenocarcinomas from a wide range of sites was studied with a panel of monoclonal antibodies with specificity for carcinoembryonic antigen, carcinoembryonic antigen with non-specific cross reacting antigen co-specificity, epithelial membrane antigen, OC 125, and OC 19.9. A range of reactivity was seen in tumours from most sites. No distinctive results were identified for a particular site of origin. Some patterns of consistent positivity or negativity for the panel of antisera used were seen, however, which if applied to metastatic deposits have the potential to assist prediction of the site of origin. These results also imply that immunohistological reactivity should be considered in the selection of a tumour marker for serological monitoring.