Among the key cell cycle regulators, cyclin D1 has been implicated most strongly in oncogenesis. This G1 cyclin is a putative proto-oncogene whose clonal rearrangement and/or amplification and mRNA overexpression occurs in several types of human neoplasias. We have now raised a series of monoclonal antibodies to human cyclin D1 and analysed its regulation at the protein level in 40 human tumour cell lines. We found that 12 cell lines displayed low or undetectable cyclin D1 protein level, while the remaining lines accumulated the protein to a level comparable to, or moderately higher than, that of four normal diploid non-immortalized cell types. The cell cycle-dependent oscillation and subcellular localization of cyclin D1 were similar in both tumour and normal cells. The protein localized to the nucleus of G1 cells, and it was reduced to immunocytochemically undetectable level in DNA-replicating cells. At the functional level, microinjection and electroporation of anti-D1 antibodies revealed that in most tumour cell lines studied, including those with amplification at the cyclin D1 locus, this cyclin is essential for cell cycle progression in G1. Some tumours, however, seem to have evolved mechanism(s) that enable them to bypass the requirement for functional cyclin D1.
- MeSH
- buněčný cyklus * MeSH
- cyklin D1 MeSH
- cykliny antagonisté a inhibitory imunologie metabolismus MeSH
- imunohistochemie MeSH
- kultivované buňky MeSH
- lidé MeSH
- monoklonální protilátky MeSH
- nádorové buňky kultivované MeSH
- onkogenní proteiny antagonisté a inhibitory imunologie metabolismus MeSH
- subcelulární frakce metabolismus MeSH
- Check Tag
- lidé MeSH
- MeSH
- histologické techniky MeSH
- imunohistochemie MeSH
- lidé MeSH
- monoklonální protilátky MeSH
- myši MeSH
- nádorový supresorový protein p53 analýza imunologie MeSH
- nádory chemie MeSH
- rekombinantní proteiny imunologie MeSH
- specificita protilátek MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
Expression of the p53 oncoprotein was examined in a wide range of primary human testicular germ-cell tumours using a new mouse monoclonal antibody (MAb) BP53-11 raised and characterized in this study, in parallel with a polyclonal rabbit antiserum CM-1. Immunohistochemistry on paraffin sections showed positive nuclear reaction in at least a fraction of malignant cells in 90 (84%) out of 107 cases studied. Aberrant accumulation of the p53 protein was found among testicular tumours of all major histological types, although generally a higher percentage of positive cases and a higher proportion of p53 over-expressing nuclei within individual lesions was observed in embryonal carcinomas when compared with seminomas. The typical heterogeneous staining pattern characteristic of histological specimens was also found in a cultured cell line derived from a human embryonal carcinoma. In contrast to immunohistochemically undetectable levels in normal testes and morphologically normal tissue areas in the tumour-bearing testes, the accumulation of the p53 protein was clearly identified in a high proportion (59% of cases) of the pre-invasive lesions with positive atypical intratubular germ cells often found in the tissue adjacent to invasive tumours. Altered expression of the p53 protein is therefore a unifying feature of the majority of invasive male germ-cell tumours and the change resulting in high levels of p53 appears to be a relatively early step in the human testicular cancer pathogenesis.
- MeSH
- choriokarcinom chemie MeSH
- dysgerminom chemie MeSH
- germinální a embryonální nádory chemie MeSH
- lidé MeSH
- mezonefrom chemie MeSH
- monoklonální protilátky diagnostické užití MeSH
- mutace * genetika MeSH
- nádorový supresorový protein p53 * analýza genetika MeSH
- teratom chemie MeSH
- testikulární nádory * chemie MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- práce podpořená grantem MeSH
Accumulation of the p53 protein was analysed in 212 human malignant lesions. Immunohistochemical staining with new polyclonal (CM-1) and monoclonal antibodies (BP 53-12 and BP53-24) to p53 on methacarn-fixed paraffin sections showed positive staining in 161 (76%). The positive tumours were found across a wide range of human malignancies including breast, colon, stomach, bladder and testis carcinomas, soft-tissue sarcomas and melanomas. The staining was always confined to the malignant lesion. Immunoprecipitation and quantitative ELISA assays established that the positive staining was associated with accumulation of the protein and that the protein was frequently in a mutant conformation. Accumulation of mutant p53 protein is therefore a common feature of human malignant disease.
- MeSH
- ELISA MeSH
- exprese genu MeSH
- geny p53 * MeSH
- imunohistochemie MeSH
- lidé MeSH
- monoklonální protilátky MeSH
- mutace * MeSH
- nádorový supresorový protein p53 analýza genetika MeSH
- nádory * genetika patologie MeSH
- protilátky MeSH
- transfekce MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- práce podpořená grantem MeSH
When defined in terms of markers for normal cell lineages, most invasive breast cancer cells correspond to the phenotype of the common luminal epithelial cell found in the terminal ductal lobular units. Luminal epithelial cells cultured from milk, which have limited proliferative potential, have now been immortalized by introducing the gene encoding simian virus 40 large tumor (T) antigen. Infection with a recombinant retrovirus proved to be 50-100 times more efficient than calcium phosphate transfection, and of the 17 cell lines isolated, only 5 passed through a crisis period as characterized by cessation of growth. When characterized by immunohistochemical staining with monoclonal antibodies, 14 lines showed features of luminal epithelial cells and of these, 7 resembled the common luminal epithelial cell type in the profile of keratins expressed. These cells express keratins 7, 8, 18, and 19 homogeneously and do not express keratin 14 or vimentin; a polymorphic epithelial mucin produced in vivo by luminal cells is expressed heterogeneously and the pattern of fibronectin staining is punctate. Although the cell lines have a reduced requirement for added growth factors, they do not grow in agar or produce tumors in the nude mouse. When the v-Ha-ras oncogene was introduced into two of the cell lines by using a recombinant retrovirus, most of the selected clones senesced, but one entered crisis and emerged after 3 months as a tumorigenic cell line.
- MeSH
- antigeny transformující polyomavirové * genetika MeSH
- buněčné dělení MeSH
- buněčné linie MeSH
- DNA virů genetika MeSH
- epitelové buňky MeSH
- fluorescenční protilátková technika MeSH
- genetické vektory MeSH
- keratiny metabolismus MeSH
- lidé MeSH
- mateřské mléko * cytologie MeSH
- myši nahé MeSH
- myši MeSH
- onkogenní protein p21(ras) genetika MeSH
- opičí virus SV40 genetika MeSH
- prsy * cytologie MeSH
- Retroviridae genetika MeSH
- Southernův blotting MeSH
- techniky in vitro MeSH
- virová transformace buněk * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
A series of 14 new mouse monoclonal antibodies (MAbs) to keratins is described and the data suggesting their potential value in the differential diagnosis of human tumours are reported. The specificities of individual MAbs of the 'C-series' presented here range from monospecificity for keratin No. 7 (MAbs C-18, C-35, C-62, and C-68), keratin No. 8 (MAbs C-15, C-43, and C-15), and keratin No. 18 (MAbs C-04 and C-08) up to the broadly reacting 'pan-keratin' MAb C-11, with the target epitopes of the remaining four MAbs being shared by different pairs of keratin polypeptides. The results of the biochemical characterization of the MAbs, together with their immunohistochemical staining patterns on frozen as well as on paraffin sections of normal human tissues, suggest that they represent a significant contribution to the growing list of anti-keratin MAbs applicable in both research and routine diagnostic pathology. The immunohistochemical examination of a wide range of human neoplasms with the new MAbs not only confirmed their value in making distinctions between carcinomas, on the one hand, and lymphomas, and gliomas, on the other, but also verified the possibility of more subtle subdivisions within the group of adenocarcinomas and their metastases. Furthermore, the identification of small subsets of breast carcinomas with decreased levels or apparent loss of the keratin No. 7 polypeptide and some cases of stomach carcinoma with apparently induced expression of this keratin suggests that such 'exceptions' must be considered when using keratin spectra as one of the criteria in differential diagnosis.
- MeSH
- 2D gelová elektroforéza MeSH
- buněčné linie MeSH
- epitel imunologie MeSH
- fluorescenční protilátková technika MeSH
- imunoblotting MeSH
- keratiny analýza imunologie MeSH
- lidé MeSH
- monoklonální protilátky diagnostické užití imunologie MeSH
- myši MeSH
- nádory chemie imunologie patologie MeSH
- specificita protilátek MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
