Histochemical and immunohistochemical characterization of surgically resected and heterotransplanted Wilms' tumor.

Nine surgically resected Wilms' tumors (WIT) and nude mouse heterotransplants from one WIT were studied by histochemistry and immunohistochemistry. Histochemistry showed acid phosphatase in all cells, while alkaline phosphatase and gamma-glutamyl transpeptidase were present in only some tubules. Using immunohistochemistry, antibodies to the intermediate filaments cytokeratin and vimentin distinguished tubular epithelium and mesenchyme, respectively. WIT tubules were also identified using antibody against a structural component (epithelial membrane antigen) and a secretory product (uromucoid) associated with distal convoluted tubules of normal kidney. Basement membrane surrounding the tubules of WIT was demonstrated using antibody to type IV collagen plus laminin. Different blastema subpopulations were negative or stained positively with antibodies to cytokeratin and vimentin. Production of basement membrane by blastema was also shown. Fetal antigen expression in WIT was examined using the monoclonal PI 153/3 and J5 antibodies. The blastema and tubules of WIT were strongly stained by PI 153/3, which did not label normal adult kidney, and weakly stained by J5, which strongly labeled glomeruli and proximal convoluted tubules of normal kidney. These studies show that WIT blastema is heterogeneous in intermediate filament subtypes, while WIT tubules more closely resemble distal than proximal convoluted tubules of adult kidneys but also retain expression of fetal antigens.

The relationship of polysialic acid and the neural cell adhesion molecule N-CAM in Wilms tumor and their subcellular distributions.

In previous studies we have reported that polysialic acid is an oncodevelopmental antigen in human kidney but its relationship to the neural cell adhesion molecule (N-CAM) remained undefined. In the present study, we showed by the combination of immunoprecipitation and immunoblotting that renal polysialic acid is a structural component of N-CAM polypeptide and that two highly sialylated N-CAM isoforms of approximately 120 kDa and 140 kDa existed in Wilms tumor. The presence of a cell surface coat composed of polysialic acid and N-CAM was revealed by immunoelectron microscopy, and morphological evidence for its involvement in modulating cell-cell adhesion has been provided. Furthermore, highly sialylated N-CAM was detectable extracellularly. N-CAM immunolabeling was present in compartments from the nuclear envelope to the plasma membrane. However, polysialic acid was only detectable at the cell surface suggesting that in Wilms tumor cells sialyl polymer synthesis may occur partially or exclusively at this site.

Localization of myoglobin in normal and neoplastic human skeletal muscle cells using an immunoperoxidase method.

Using an immunoperoxidase method, myoglobin is localized in the cytoplasm of normal and neoplastic human skeletal muscle. The staining intensity is variable in individual cells. This can be explained by the variable concentration of myoglobin in the different fiber types of normal muscle or the different degree of differentiation in neoplastic muscle cells, respectively. A potential application of this method in tumor pathology is discussed in connection with myoglobin as a tissue-specific marker for skeletal muscle neoplasms.

Renin messenger RNA localization in congenital mesoblastic nephroma using in situ hybridization.

Renin messenger (m) RNA distribution was studied in congenital mesoblastic nephroma, a usually benign renal tumour of early infancy which may be associated with excess renin production and hypertension. Using in situ hybridization with synthetic radiolabelled oligonucleotide probes combined with immunohistochemical studies, renin expression was found in areas of tumours containing recognizable cortical structures including glomeruli and tubules. Renin mRNA was also detected in vessels and larger vascular spaces within the tumour not associated with cortical structures. Cells in the tumour vessel walls and sinusoids which expressed renin also stained positively for vascular smooth muscle-specific alpha actin.

Renin in mesoblastic nephroma: an immunohistochemical study.

Congenital mesoblastic nephroma (CMN) is the most common renal tumor of early infancy. It is usually evident at birth as an abdominal mass and is composed of spindle-shaped cells resembling smooth muscle cells and fibroblasts. There is macroscopic and microscopic infiltration of the surrounding kidney and entrapped tubules and glomeruli are common at the edge of the tumor. In this report, we describe a case of CMN associated with hyperreninaemia and hypertension. We examined the tumor from this case and 11 other cases for the presence of renin using a polyclonal antibody to human kidney renin. Hypertension was present in three of four additional cases for which records were available. Immunoreactive renin was present in ten of the 12 cases studied. In all of these cases, intense staining was present in vessels within the areas of the trapped cortex. In seven cases, renin was identified in the walls of vessels within the tumor itself without identifiable adjacent cortical structures. These findings indicate that CMN may often be associated with primary reninism in early infancy.

Extrarenal Wilms' tumor: an analysis of four cases.

During a review of Wilms' tumor, four located external to the kidney were identified. Patient age ranged from 7 months to 4 years; three were female. One neoplasm was located in the parametrial connective tissue to the left of the uterus; both kidneys were radiographically normal. Three neoplasms were located in the right retroperitoneum adherent to the surface of the ipsilateral kidney, but separated from the parenchyma by a thick fibrous capsule. Two were attached to the upper pole, while the third was attached to the midportion of the kidney. Radiologic studies showed displacement of all three kidneys, but intravenous pyelogram (IVP) revealed no calyceal distortion. All four neoplasms were favorable histology Wilms' tumor: one was monophasic epithelial type, one was monophasic blastemal type, and two had a mixture of stromal, epithelial, and blastemal tissue. No teratomatous elements were present. Immunoperoxidase staining for cytokeratin (AE-1, AE-3, CAM 5.2), vimentin, and epithelial membrane antigen (EMA) showed the strongest focal positive staining of tubular structures with CAM 5.2, and slight staining with EMA. Staining reaction to vimentin was variable, but negative in most areas. Three tumors extracted from paraffin were diploid by quantitative flow cytometric DNA analysis; in one case, flow cytometry could not be performed. Clinical follow-up from 2 years to 6 years showed all children to be alive without evidence of disease. Based on the similarity to conventional renal Wilms' tumor, these findings support the hypothesis of displaced mesonephric/metanephric rests for the origin of extrarenal Wilms' tumor.

Cell differentiation in Wilms' tumor (nephroblastoma): an immunohistochemical study.

Using an indirect immunoperoxidase technique, we tested frozen specimens from 12 Wilms' tumors with monoclonal antibodies (MoAbs) reacting against a large panel of molecules including laminin, fibronectin, cytokeratin, vimentin, villin, CD24, CALLA/CD10, CR1, CD26, class I and class II major histocompatibility complex (MHC) molecules, and endothelium factor VIII. These molecules were chosen because they are markers of specific segments of the mature kidney and because their loss or acquisition is indicative of different steps of human nephrogenesis. KI67 MoAb was used to evaluate the proliferating activity of the cells. The blastemal component (cell compact areas) of Wilms' tumors consisted of vimentin-positive cells with a fibronectin network. However, signs of epithelial maturation were present in compact areas where cytokeratin-positive cells producing laminin were observed. The cells exhibited a high degree of proliferating activity. The tubule formations consisted of cytokeratin-positive cells and had a defined laminin border. All the cells, whether in compact areas or in tubules, were strongly CD24-positive. Some tubular formations showed signs of proximal maturation with the presence of CALLA, CD26, and even villin. In four cases class I-MHC molecules were expressed by some tubular cells. Large cystic cavities present in five cases were edged by cytokeratin, CD24-positive cells, or by vimentin, CALLA, CR1-positive cells. Some glomeruloid bodies, present in two cases, were also composed of vimentin, CALLA, and CR1-positive cells which correspond to the mature podocyte phenotype. The interstitial tissue contained mainly laminin and fibronectin network with macrophages and few CD3 lymphocytes. The presence of large cells with muscular differentiation was noted; round vimentin and CD26-positive cells were also seen. The endothelial cells of the vessels exhibited vimentin, factor VIII, and class I and class II MHC molecules as do mature cells, but in some cases the endothelial cells lacked class II molecule expression and were CALLA-positive. These results which confirmed and extended those previously described show that cell differentiation in Wilms' tumor mimics that observed during metanephros development. Moreover, this study shows that tumoral cells in nephroblastoma share several antigens with cells from lymphoid lineage (CD24, CALLA, and CD26) as do developing and mature kidney cells. Such cell phenotype dissection provides a useful and reliable tool for testing the influence of various factors on the development of hetero-transplanted or cultured Wilms' tumors.

Lectin histochemistry of Wilms' tumor. Comparison with normal adult and fetal kidney.

The lectin histochemical staining patterns of nine surgically resected Wilms' tumors (WIT) (five classical, one rhabdomyomatous, one monomorphic tubular, and two blastematous) and four WIT heterotransplants in nude mice were compared with those of six normal adult and fetal kidneys using 11 biotinylated and fluorochrome-labeled lectins representing a spectrum of sugar specificities. Tubular epithelial cells of fetal and adult kidney demonstrated a complex pattern of glycosubstances, as detected by staining with most of the lectins. The dysplastic tubules of WIT and WIT heterotransplants resembled distal convoluted tubules and collecting ducts of adult kidney, as detected by staining with lectins that reacted exclusively with these portions of the nephron. A simpler lectin affinity profile was observed with blastema of normal fetal kidney and blastema of WIT and WIT heterotransplants. Thus, fetal, adult, and tumor tubules and fetal and tumor blastema resemble each other in terms of lectin binding patterns.

Histochemical evidence for tubule segmentation in a case of Wilms' tumor.

A lectin histochemical and immunohistochemical approach was used to compare the different histologic elements of an unusual case of Wilms' tumor with normal kidneys. This case was stained with ten lectin-horseradish peroxidase conjugates; immuno-stained for Tamm-Horsfall glycoprotein, epidermal keratin, and vimentin; and compared with 19 control kidneys. The morphologic and cytochemical properties of various tumor elements in this specimen served to identify them as tumor analogs of all segments of the normal kidney tubules except distal tubules. Evidence for Wilms' tumor differentiation is provided by this case, whose epithelium histochemically resembles normal human epithelial cells.

Mesoblastic nephroma contains fibronectin but lacks laminin.

Non-metastatic mesoblastic nephromas from four young children were shown to contain fibronectin but not laminin using an immunoperoxidase staining procedure. In contrast, one metastasising spindle celled sarcomatous tumour from a neonate was laminin positive. During embryogenesis primitive nephrogenic mesenchyme contains only fibronectin and no laminin; metanephric blastema (permanent kidney) is positive for laminin. The staining for fibronectin and laminin may help to ascertain the histogenesis of different types of renal tumour.

Feulgen DNA values in Wilms' tumour in relation to prognosis.

Single-cell DNA measurements were performed on Feulgen-stained nuclei in 4 mu tumour sections from 30 patients with Wilms' tumour, all treated with initial nephrectomy. Fifteen patients died from their tumours within 2 years of nephrectomy. The remaining 15 had survived for at least 10 years without evidence of recurrence. Based on the histograms, the tumours were classified as being euploid (E type) or aneuploid (A type). Twenty tumours were euploid and 10 aneuploid. The euploid tumours predominated in the group of survivors. The group of 10 aneuploid tumours contained 7 tumour deaths. The tendency for aneuploid tumours to be associated with early tumour death, however, was not statistically significant.

Glycosaminoglycan in the blood and renal tissue of a patient with nephroblastomatosis.

Bilateral nephroblastomatosis was diagnosed in a 15-month-old white female. Prior to surgery, multiple peripheral blood smears (Wrights' stain) revealed an azurophilic staining extracellular material. When serum was added to a three percent acetic acid solution, a floccular, fibrous precipitate formed at the meniscus of the tube. Serum protein electrophoresis on cellulose acetate support media resulted in a distorted pattern which corrected to a normal pattern upon treatment with hyaluronidase. These peripheral blood abnormalities disappeared following a left nephrectomy. Quantitative chemical analysis of diseased renal tissue yielded 81 micrograms of readily extracted glycosaminoglycan (GAG) per gram of tissue. The importance of abnormal glycosaminoglycan production in patients with malignant disease is discussed both in terms of clinical importance and possible roles of cell exudates.

Nephroblastoma. A comparative immunocytochemical and lectin-histochemical study.

Nephroblastomas (Wilms' tumors) from a dog, a bird, a pig, and a child were subjected to comparative immunocytochemical and lectin-histochemical analysis along with normal renal tissues from the same animals. Primary rabbit and mouse anti-human antibodies directed at intermediate-filament proteins, neuron-specific enolase, S100 protein, and epithelial membrane antigen were employed, as were biotinylated peanut agglutinin, soybean agglutinin, wheat germ agglutinin, and lectins from Dolichos biflorus and Ulex europaeus. The human neoplasm showed positivity for cytokeratin and epithelial membrane antigen and bound peanut, soybean, and wheat germ agglutinins in epithelial areas. Among the animal tumors, the porcine and canine nephroblastomas showed immunoreactivity for cytokeratin and vimentin in epithelial and blastematous areas, respectively. In addition, they were positive for S100 protein in epithelial foci. All of these results were substantiated by staining patterns in nonhuman kidneys. None of the neoplasms or renal tissues showed reactivity to the other antigens that were assessed. In the porcine tumor, endothelial cells bound D biflorus lectin, and epithelial areas were stained by U europaeus lectin. The avian nephroblastoma bound peanut, soybean, and wheat germ agglutinins, while the canine neoplasm showed no lectin-histochemical reactivity. These data appear to reflect limited immunohistological and histochemical similarities between nephroblastomas of different vertebrates.

Neuron specific enolase: a marker for differential diagnosis of neuroblastoma and Wilms' tumor.

Thirteen human neuroblastoma and six Wilms' tumor biopsies have been analyzed for neuron specific enolase (NSE). THe relative activity of NSE in the neuroblastomas (including ganglioneuroblastoma and ganglioneuroma) ranged from 28% to 62.5% of total enolase activity. The corresponding figures for the Wilms' tumors were 1% to 4.5%. It appears that NSE can serve as a biochemical marker for neuroblastoma and be useful in the differential diagnosis of neuroblastoma and Wilms' tumor.

Nephroblastoma with neuroendocrine differentiation.

A nephroblastoma (Wilms' tumor) with morphological, histochemical, immunohistochemical and ultrastructural evidence of neuroendocrine differentiation is described. Whereas areas of neural differentiation and occasional argyrophilic cells in cases of Wilms' tumor have been previously reported, the unique characteristic in this case was the extent of the neuroendocrine differentiation, as shown by a strong Grimelius stain of over 90% of the blastematous cells. Immunoperoxidase studies employing antibodies to neuron-specific enolase and ultrastructural data were also in favor of the neuroendocrine differentiation and suggested the existence, in addition to the already reported variant of Wilms' tumor with neural differentiation, of a neuroendocrine variant which may be part of the histologic spectrum of this neoplasm.

[Flow cytometry of DNA in research on kidney tumors]. / Protochnaia tsitometriia DNK v issledovanii opukholei pochek.

The DNA content was studied in 22 patients (including 10 children) with renal tumours. Renal clear cell carcinomas were mainly diploid (5 of 6 cases); one clear cell and other variants of renal cell carcinoma were aneuploid. There was a correlation between the degree of tumour cell anaplasia and ploidy of renal cell carcinomas: all diploid carcinomas were of I and II degree of anaplasia, aneuploid carcinomas, except one case, were of degree III of anaplasia. 5 out of 9 nephroblastomas were diploid, 4, aneuploid. The distinctive features of nephroblastoma were pronounced proliferative activity of tumour cells and a low DNA index of the aneuploid cell line which in all cases was localized in the vicinity of the diploid region. The remaining tumours (papillary epithelial nephroma, juxtaglomerular cell tumour and malignant schwannoma) were diploid with a relatively low proliferative activity of tumour cells.

[A highly sialylated, embryonic form of the neural cell adhesion molecule in Wilms tumor: identification of a cell adhesion molecule as a onco-developmental antigen]. / Nachweis der hochsialylierten, embryonalen Form des Neuralen Zelladhäsionsmoleküles im Wilms Tumor: Identifizierung eines Zelladhäsionsmoleküles als onko-developmentales Antigen.

The neural cell adhesion molecule NCAM is a general Ca2+ -independent cell adhesion molecule which exerts important functions during the development of the nervous system. NCAM polypeptides exist in various isoforms all of which have a similar extracellular domain structure containing a homophilic binding site for establishment of cell-cell contacts. A common structural feature of NCAM is the presence of homopolymers of alpha 2,8-linked sialic acid residues, the so-called polysialic acid, which is developmentally regulated. It undergoes a change from a highly less sialylated short chain form from embryonic to adult life. The polysialic acid regulates the homophilic adhesive properties of NCAM. The highly sialylated form of NCAM typically found in developing tissues decreases homophilic NCAM-NCAM interactions and, therefore, cell-cell contacts due to its large excluded volume and electric repulsive forces. We have used a monoclonal antibody against polysialic acid, polyclonal antibodies against NCAM, polysialic acid specific bacteriophage-encoded endoneuraminidases and a NCAM cDNA to investigate this molecule by light and electron microscopic immunolabeling, in situ hybridization and immunochemistry. The highly sialylated form of NCAM was found to be expressed in a developmentally-regulated fashion in embryonic kidney, undetectable in the normal adult kidney and re-expressed in Wilms tumor. In Wilms tumor the blastemal cell masses as well as epithelial differentiations such as tubules and glomeruloid bodies were immunolabeled whereas the stroma did not label for polysialic acid. Combination of immunoprecipitation and immunoblotting using antibodies against polysialic acid and NCAM, respectively, directly demonstrated structural relationship of renal polysialic acid with NCAM polypeptide. By immunoblot analysis two NCAM isoforms of approximately 120 and 140 kD were found in Wilms tumor. Immuno-electron microscopy provided direct morphological evidence for prevention of cell-cell contacts due to the presence of a thick cell surface coat composed of polysialic acid. In nephroblastomatosis complexes only blastemal cells exhibited immunocytochemically detectable polysialic acid. All other investigated kidney tumors, i.e. clear cell sarcoma, malignant rhabdoid tumor, cystic nephroma and renal cell carcinoma, were negative. These data allow the conclusion that the highly sialyated, embryonic form of NCAM is an onco-developmental antigen in human kidney. At the same time this is the first observation that a molecule with a well defined role for controlled cell migration and differentiation during embryonic organ development represents an onco-developmental antigen.