Michael Altmannsberger

Diagnosis of human childhood rhabdomyosarcoma by antibodies to desmin, the structural protein of muscle specific intermediate filaments

SummaryFour embryonal rhabdomyosarcomas, one tumor diagnosed as an undifferentiated sarcoma, probably a rhabdomyosarcoma, and six different non-muscular sarcomas were investigated with antibodies specific for different intermediate filament types. The tumor cells in the rhabdomyosarcomas and the undifferentiated tumor were stained clearly by antibodies to desmin, the intermediate filament type characteristic of muscle. The staining of tumor cells by antibodies to vimentin, the intermediate filament type characteristic of certain cell types of mesenchymal origin including myoblasts, was different in these 5 cases. In one case of embryonal rhabdomyosarcoma nearly all tumor cells were stained, but in the remaining cases few or no tumor cells were positive with the vimentin antibody. In these rhabdomyosarcomas not only the large rhabdomyoblasts, but also the small undifferentiated cells were labeled by antibodies to desmin. In the latter cell type the desmin filaments were arranged typically in coils.In contrast, tumor cells in the non-muscular mesenchymal sarcomas were stained only by antibodies to vimentin but not by antibodies to desmin or prekeratin. The retention of the desmin marker characteristic of normal muscle in cases of rhabdomyosarcoma not only allowed the undifferentiated desmin-positive sarcoma to be classified as rhabdomyosarcoma but also suggests that the use of antibodies to desmin could be very helpful in the future for the diagnosis of undifferentiated rhabdomyosarcomas.

The distribution of keratin type intermediate filaments in human breast cancer: An immunohistological study

SummaryAntibodies to different intermediate filament proteins can be used to distinguish cells of epithelial, mesenchymal, muscle, glial and neuronal origin. Antibodies to prekeratin which characterize cells of epithelial origin, and antibodies to vimentin which recognize cells of mesenchymal origin have been used to study twenty cases of breast carcinoma (sixteen infiltrating ductal carcinomas and four infiltrating intraductal carcinomas), two cases of cystic breast disease, two fibroadenomas and one case of benign cystosarcoma phylloides. The prekeratin and vimentin were detected using specific antibodies to these proteins by immunofluorescence microscopy using alcohol fixed paraffin-embedded tissues. In eighteen out of the twenty carcinomas the tumor cells were strongly and specifically stained by antibodies to prekeratin. Different tumors gave different patterns of prekeratin staining. In contrast, when the same specimens were tested with the vimentin antibody, the tumor cells were unstained, and instead only the usual strong staining to fibroblasts and blood vessels in the stroma was observed. In cystic breast disease, fibroadenomas, and benign cystosarcoma phylloides, cells of epithelial origin were strongly stained by the prekeratin but not by the vimentin antibody.

Various sympathetic derived human tumors differ in neurofilament expression. Use in diagnosis of neuroblastoma, ganglioneuroblastoma and pheochromocytoma.

SummaryWe have extended our analysis of human tumors using antibodies specific for each of the five types of intermediate filaments to neuroblastoma, ganglioneuroblastoma, pheochromocytoma, ependymoblastoma, and alveolar soft part sarcoma. Tumor cells in the three cases of neuroblastoma, as well as in the single case of alveolar soft part sarcoma, did not react positively with sera directed against any of the five intermediate filament types. We suppose, therefore, that neuroblastoma at least may be derived from a cell type — possibly present in peripheral neurones — whichin vivo has very few or no intermediate filaments. In ganglioneuroblastoma and in pheochromocytoma the tumor cells were positive when tested with antibodies directed against neurofilaments and negative with those directed against other intermediate filament types. The ependymoblastoma was positive when tested with antibodies directed against glial fibrillary acidic protein (GFA) and negative when tested with antibodies against other intermediate filament types. Use of antibodies to the different intermediate filament types appears to be a valid way in which to classify tumors, and so far the data presented here and elsewhere support the hypothesis that tumor cells retain the intermediate filament type typical of their cell of origin. Wider use of these sera would seem particularly useful in cases such as neuroblastoma, rhabdomyosarcoma or lymphoma where diagnosis is currently difficult using conventional histological stains.

Keratin polypeptide distribution in benign and malignant breast tumors: subdivision of ductal carcinomas using monoclonal antibodies.

SummaryMonoclonal antibodies which recognize one or only a few keratin polypeptides have been used to study the distribution of different keratins in benign and malignant breast lesions by immunocytochemical methods. Seven monoclonal antibodies which recognized either different keratin polypeptides by immunoblotting techniques, or identified different epithelial cell types in complex tissues were used. In two mastopathies and three fibroadenomas the antibody 1u5 stained luminal cells as well as myoepithelial cells. In contrast the antibodies CK7, Troma 1, CK2 and KA4 labeled only luminal cells, whereas antibody CKB1 decorated only myoepithelial cells. All 15 ductal carcinomas showed a uniform staining of tumor cells with the antibodies Troma 1, CK2, KA4 and 1u5. The antibody CK7 also stained all ductal carcinomas, but in two specimens the staining was heterogeneous. The antibody CKB1 decorated only the preexisting myoepithelial cells in 11 of 12 ductal carcinomas but in the remaining specimen the tumor cells were also strongly positive. Tumor cells in lobular carcinomas were labeled by antibodies CK7, Troma 1, CK2, KA4, but not by CKB1. The antibody CKS1 showed no staining of any of the benign and malignant breast lesions.

Distinction of nephroblastomas from other childhood tumors using antibodies to intermediate filaments

SummaryTen nephroblastomas were investigated by antibodies to inter-mediate filaments. In seven cases, which in light microscopy were characterized by the presence of blastema and tubules, immunofluorescence microscopy with IF-specific antibodies reveals expression of cytokeratin and vimentin in blastema cells, while tubules were only labelled by the cytokeratin antibodies. This result was independent of whether the conventional cytokeratin antibody or monoclonal antibodies specific for cytokeratin 18 were used. Stroma cells were vimentin-positive. In two cases nephroblastomas were undifferentiated and also lacked tubuli formation. In both these tumors blastema cells were vimentin-positive and cytokeratin-negative. Finally one case of clear cell sarcoma of the kidney could only be labelled by the vimentin antibody. Thus antibodies to intermediate filaments seem to be useful tools to distinguish nephroblastomas from neuroblastomas or rhabdomyosarcomas, especially in cases of metastasis.

Diagnostic value of intermediate filament antibodies in clinical cytology

SummaryAntibodies to intermediate filament (IF) proteins can distinguish the major tumor groups as shown by results with sectioned human material. In this study we evaluate the use of similar methods in the cytology of human tumors. Smears obtained from fine needle aspiration biopsies were investigated using well characterized antibodies, each specific for only one of the five types of intermediate filaments. Tumor cells of different carcinomas, thymomas, and the epithelial part of pulmonary blastomas were positive with antibodies recognizing cytokeratins. Tumor cells in non-muscle sarcomas, including lymphoma and Ewings sarcoma, could be specifically identified with antibodies to vimentin. Tumor cells of muscle sarcomas were desmin-positive. Finally, tumor cells in pheochromocytoma and bronchus carcinoid were positive with antibodies specific for neurofilaments. Specimens were also examined in parallel using conventional cytochemical stains, such as May-Grünwald-Giemsa. In addition, in most cases sections of the tumor were examined both by histology and IF typing of frozen sections to confirm the diagnosis made on the cytologic specimens. The results show that IF typing is a valuable diagnostic aid in clinical cytology.

Identification of Actin Microfilaments in the Intracytoplasmic Inclusions Present in Recurring Infantile Digital Fibromatosis (Reye Tumor)

The diagnostic intracytoplasmic perinuclear inclusion bodies within the fibroblastic tumor cells of recurring digital fibrous tumor of childhood (Reye tumor) were found to be ultrastructurally composed of condensed microfilaments that were continuous with axially oriented cytoplasmic filament bundles running toward the cell membrane. Immunofluorescence microscopy performed with antibodies raised against vimentin and against actin revealed a strong positive reaction for vimentin in the tumor cell cytoplasm, whereas the spherical inclusion bodies were actin-positive. From these results the following conclusions may be drawn: (I) the mesenchymal origin of the Reye tumor cells is consistent with the identification of vimentin intermediate filaments and (II) the actin-positive spherical inclusion bodies appear to represent pathological aggregations of microfilaments.

Esthesioneuroblastoma: Ultrastructural, immunohistological and biochemical investigation of one case

SummaryA case of esthesioneuroblastoma, the pathological diagnosis of which almost always causes great difficulties, was investigated ultrastructurally, biochemically, and immunohistologically, using antibodies against the five known types of intermediate filaments [keratin, vimentin, desmin, glial fibrillary acidic protein (GFAP) and neurofilaments]. The tumour cells did not react with antibodies against any of the five intermediate filament proteins. Ultrastructural investigations showed dense cored secretory granules in the cytoplasm and cell processes. Thus, immunohistology offers by “exclusion” a differential diagnosis to avoid often misdiagnosed tumours (undifferentiated carcinomas, embryonal rhabdomyosarcomas, and malignant lymphomas), since carcinomas react with antikeratin, embryonal rhabdomyosarcomas with antibodies to desmin and malignant lymphomas show immunofluorescence with antibodies to vimentin. The biological behaviour (age distribution, tendency to metastasize), the normal values of biochemical parameters, homovanillic acid and vanilmandelic acid (HVA, VMA), and the absence of neurofilaments distinguish this type of tumour from the peripheral sympathetic neuroblastoma.

Chemically Induced Esthesioneuroepithelioma: Ultrastructural Findings

Tumors of the olfactory epithelium of rats were induced with two different nitrosamines: 2,6-dimethylnitrosomorpholine and N-nitrosopiperidine. Both carcinogens yielded identical tumors consisting of small, undifferentiated, neuroblastic cell elements without specialized cell contact. Cell processes contained microtubuli, centrioles, and neurosecretory granules. Two kinds of rosettes were encountered frequently: Neuroblastic Homer Wright rosettes consisted of undifferentiated cells, surrounding a minute lumen filled with amorphous material; and Flexner rosettes showed a higher degree of maturation. Inside their central lumen, cell processes with characteristic features of olfactory sensory cells (basal bodies, cilia, centrioles, microtubuli) could be demonstrated. The stem cell of this tumor is most likely the undifferentiated light basal cell inside the olfactory epithelium, since its ultrastructural appearance and its cytoskeleton are alike. At least under neoplastic conditions, this stem cell may likewise differentiate into epithelial cells, since transition to squamous cell carcinomas has been observed. In view of their overwhelming similarity to their human counterpart, the induced tumors are most likely to represent esthesioneuroepitheliomas.