J. J. G. Puts

Antibodies to intermediate filament proteins in the immunohistochemical identification of human tumours: an overview

SummaryIntermediate-sized filament proteins (IFP) are tissue specific in that antibodies to keratin, vimentin, desmin, glial fibrillary acidic protein (GFAP) and the neurofilament proteins can distinguish between cells of epithelial and mesenchymal origin as well as of myogenic and neural origin respectively. Malignant cells retain their tissue-specific IFP, which makes it possible to use these antibodies in tumour diagnosis. Carcinomas are exclusively detected by antibodies to keratin. Monoclonal antibodies to keratin have allowed the differentiation between subgroups of epithelial tumours until now between adenocarcinomas and squamous cell carcinomas. Lymphomas, melanomas and several soft tissue tumours are distinctly recognized by antibodies to vimentin. On the other hand, rhabdomyosarcomas and leiomyosarcomas are positive for desmin, while astrocytomas give a strong reaction with GFAP antibodies. Thus, antibodies to IFP are useful tools for differential diagnosis in surgical pathology.

Expression of cytokeratins in early neoplastic epithelial lesions of the uterine cervix

SummaryPolyclonal and monoclonal antibodies to cytokeratin polypeptides were used to study the expression of these intermediate filament proteins in normal, squamous metaplastic, and neoplastic epithelium of the uterine cervix, in order to investigate the morphogenesis of early epithelial changes preceding cervical squamous cell carcinoma. A polyclonal keratin antiserum showed a positive reaction in all different epithelial cell types of the uterine cervix. A positive reaction was also found in subcolumnar reserve cell hyperplasia, in squamous metaplastic and dysplastic cells, and in (squamous) carcinoma in situ. A monoclonal antibody specific for columnar epithelium (RGE 53) gave a positive reaction in endocervical columnar cells and in some immature metaplastic cells but was negative in subcolumnar reserve cells, squamous (metaplastic} cells, dysplastic cells, and most cases of carcinoma in situ. Another monoclonal cytokeratin antibody (RKSE 60) pointed to early keratinization in light microscopically nonkeratinizing squamous (metaplastic) and dysplastic epithelium. A possible overlap in staining patterns of RGE 53 and RKSE 60 was seen in some cases of immature metaplasia. Morphologic changes occurring in the transformation zone upon dedifferentiation are accompanied by alterations in cytokeratin expression. Similarities in cytokeratin expression were found between dysplasia and carcinoma in situ on one hand and subcolumnar reserve cell hyperplasia and squamous metaplasia on the other. This study favors an epithelial origin and a squamoid nature of subcolumnar reserve cells.

Differential diagnosis of human carcinomas, sarcomas and their metastases using antibodies to intermediate-sized filaments☆

Intermediate filaments (IF) are tissue-specific in so far that epithelial, mesenchymal, muscle and neural tissue types can be distinguished by the use of specific antibodies to keratin, vimentin, desmin and neurofilaments or glial filaments respectively. We have examined the possibility of using these sera in the differential diagnosis of human malignant tumors. Using antisera to human skin keratin and bovine lens vimentin we could differentiate between carcinomas (keratin +) and sarcomas (vimentin +). Furthermore, we could show that when cells become malignant and metastasize they retain their original IF and do not develop additional IF systems. We conclude that antibodies to IF proteins are powerful tools in the hands of a pathologist as an additional method to improve identification of tumors and their metastases.

Application of antibodies to intermediate filament proteins in simple and complex tumors of the female genital tract

Polyclonal antibodies to cytokeratins, vimentin, and desmin and monoclonal antibodies to vimentin and to individual cytokeratin polypeptides, specific for glandular epithelia (RGE 53) or kertinizing stratified squamous epithelia (RKSE 60), have been applied in gynecological tumors with simple or complex composition. In general, tumors with simple composition showed reaction patterns fitting their known epithelial or mesenchymal nature, i.e., cytokeratin positivity in epithelial tumors only, vimentin positivity in mesenchymal tumors, and expression of desmin and vimentin in muscle cell tumors. Rather frequently, coexpression of cytokeratins and vimentin was noted in endometrial adenocarcinomas. Tumors with complex composition, such as müllerian mesodermal mixed tumors (MMMTs), that may pose considerable problems in conventional histopathology revealed various reaction patterns, with either expression of only cytokeratins or coexpression of cytokeratins and vimentin in carcinomatous areas and expression of only vimentin in sarcomatous areas. However, in addition, some MMMTs contained cells that were also positive for desmin. Intermediate filament protein immunohistochemistry appeared to be helpful in establishing a diagnosis of MMMT and in characterizing the different tumor components, which may prove to be useful in the evaluation of gynecological treatment protocols.

Immunohistochemical identification of Langerhans cells in normal epithelium and in epithelial lesions of the uterine cervix

Using the double label indirect immunofluorescence technique we have studied vimentin-positive cells present in normal ecto- and endocervical epithelium, subcolumnar reserve cell hyperplasia, and squamous metaplastic and dysplastic epithelium of the uterine cervix. Monoclonal antibodies to Ia- and T6-antigens were applied in the examination of the expression of these membrane markers by such cells. Our studies reveal the presence of a relatively large number of vimentin-positive and T6-positive (Langerhans) cells in normal ectocervical stratified squamous epithelium, a small number in endocervical columnar epithelium, and a larger number in subcolumnar reserve cell hyperplasia and in immature squamous metaplasia. In this respect, mature squamous metaplastic epithelium shows a great resemblance to normal ectocervical stratified squamous epithelium. In contrast with previous reports in the literature we could only demonstrate small numbers of Langerhans cells in cases of dysplasia. The clinicopathological significance of these findings is discussed.

Use of intermediate filament antibodies in the differential diagnosis of gynecological neoplasia

All eukaryotic cells contain an intracellular network of protein filaments with varying diameters, the cytoskeleton. Next to microfilaments and microtubules, lo-nanometer filaments can also be recognized. These so-called intermediate-sized filaments comprise a considerable part of the cytoskeleton and have been described to be tissue-specific (Franke et al., 1981). Recent investigations have shown that all types of epithelial cells contain cytokeratins, while mesenchymal cells contain vimentin as the protein content of their intermediate filaments (I.F.). Muscle cells contain desmin I.F., while neural tissues contain GFAP or neurofilaments (Osborn et al., 1981). Antibodies raised against these different types of I.F. proteins can be used in the immunohistochemical characterization of normal tissues as well as of benign and malignant tumors. For example, carcinomas contain only cytokeratins, while malignant lymphomas contain only vimentin (Osborn and Weber, 1983; Ramaekers et al., 1983a). Since cytokeratins consist of a number of different proteins which occur in tissue-specific combinations (Moll et al., 1982), monoclonal antibodies to these cytokeratin subsets allow distinction between different kinds of epithelia. For example, we have prepared the monoclonal antibody RGE 53 which is able to distinguish squamous from columnar epithelium (Ramaekers et al., 1983b). When applied to frozen sections of the squamo-columnar junction of the uterine cervix, for example, a polyclonal cytokeratin antibody stains both ectocervical squamous and endocervical columnar epithelium, whereas the monoclonal antibody only stains the columnar cells (Fig. 3). This antibody can also discriminate between tumors originating from these different types of epithelial cells. It shows a positive reaction in adenocarcinomas but is negative in squamous cell carcinomas. We have applied polyclonal and monoclonal antibodies to different I.F. proteins in the diagnosis of gynecological tumors. The specificity of the antibodies (see Table IV) was first tested on gynecological tumors which normally do not present problems in the differential diagnosis. In cases of papilliferous serous cystadenocarcinomas of the ovary (Fig. 4), a positive staining reaction with polyclonal cytokeratin antibodies is seen. The tumor cells are negative with the vimentin antibody, while, however, stromal elements are strongly positive for this antibody. Similar results are obtained with papilliferous mutinous cystadenocarcinomas of the ovary. Also adenocarcinomas of the uterine corpus and cervix (Fig. 5) show this positive reaction for cytokeratin only. both in

Monoclonal Antibodies to Cytokeratins in the Study of Premalignant Lesions of the Uterine Cervix

Abstract Cytokeratins are intermediate filament proteins that occur specifically in epithelial cells. Using antibodies to these proteins epithelial cells can be detected specifically by immunohistochemical methods. Thus, using a polyclonal rabbit antiserum to keratin in the indirect immunofluorescence technique on frozen tissue sections from normal cervix, ectocervical stratified squamous epithelium, endocervical columnar epithelium and the so-called reserve cells show a positive staining reaction. A monoclonal antibody, raised against cytokeratin 18 specifically recognizes columnar epithelial cells. This type of cytokeratin is not detectable in reserve cells, endocervical nature squamous metaplastic and dysplastic epithelial cells. A second monoclonal antibody specifically stains keratinizing cells in stratified squamous, squamous metaplastic and dysplastic epithelium. These data indicate that alterations in cytokeratin expression occur in premalignant lesions of the uterine cervix.