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R Dulbecco

Publications and source records attributed to R Dulbecco.

At least 37 records · Page 2Linked to original sources
PubMed1988

Stochastic development of invasive potential in rat mammary tumors induced by N-methyl-N-nitrosourea.

We have studied the progression of breast tumors induced by N-methyl-N-nitrosourea in Wistar/Furst rats during serial passages through isogeneic animals, using characteristic markers previously defined. The tumors contained various types of structure, which evolve from intraductal nodes to duct-like structures and then to invasive strands or cells. Most primary tumors were not invasive but became so in subsequent transfers. The distribution of invasive and noninvasive tumors in the various lineages was not random, implying that the tumors differed from their inception in the propensity toward invasion. In different lineages invasive structures appeared repeatedly and independently but with different frequencies, suggesting that they were produced by a stochastic mechanism with different probability of occurrence. The possible nature of this mechanism is discussed. This experimental system is useful for further studies of tumor invasion; the results obtained are relevant for understanding and evaluating the evolution of human breast cancer.

Animals↗
PubMed1988

Reversion toward an earlier stage of differentiation and loss of polarity during progression of N-methyl-N-nitrosourea-induced rat mammary tumours.

To work toward an understanding of the mechanisms of cancer progression, we have followed the changes in expression of several genes in rat mammary carcinomas during the development of invasion. Tumors were induced by N-methyl-N-nitrosourea and serially transplanted to isogeneic rats. Gene expression in individual cells was followed by immunofluorescence microscopy. We show that invading cells do not express the same genes as the primary tumor or normal adult mammary gland but are similar to the primitive mammary duct cells present in late embryos. In the most advanced tumors, the most fundamental epithelial characteristic, polarity, is lost. Relative to the mechanism of invasion, we observe that the basal lamina, often thinner than normal, always coats the strands of invading cells and may be entirely absent from single invasive cells only. These observations suggest that the basal lamina ultimately disappears because of reduced expression of the genes or the failure to assemble the components. In these tumors it is the progressive alteration in gene expression rather than the enzymatic breakdown of the basal lamina that leads to invasion. These results show that the most important aspect of cancer progression is the progressive alteration of gene expression. They also raise questions about the mechanism that causes tumor cells to retrace the steps in their differentiation.

Animals↗
PubMed1986

Organization of cytokeratin bundles by desmosomes in rat mammary cells.

In a rat mammary epithelial cell line, LA-7, cytokeratin bundles recognized in immunofluorescence by a monoclonal antibody (24B42) disappear after trypsinization of cultures and are gradually reformed after replating. We have followed the time course of cytokeratin filament reappearance by growing cells in low calcium medium (0.1 mM) which prevents desmosome formation, and then shifting to high calcium (1.8 mM) to start the process. By fixing the cells at various intervals and staining them in immunofluorescence for 24B42 cytokeratin and for desmosomal proteins, we found that cell to cell contact and desmosome formation are prerequisites for keratin filament formation in these cells. EGTA treatment, by disassembling desmosomes, causes the cytokeratin filaments to disappear and the 24B42 protein to pass into a soluble form in this cell line, as ascertained by 100,000 g fractionation and immunoenzymatic assay. Cycloheximide treatment also causes cytokeratin filaments to disappear, indicating that protein synthesis is needed for normal filament maintenance. In another related cell line (106A-10a) and in HeLa cells, trypsinization and EGTA exposure do not cause a complete loss of 24B42 immunofluorescence, although distinct filaments disappear, indicating the presence in these cells of different organizing centers, besides desmosomes, for cytokeratin bundle formation. LA7 cells therefore seem to have a cytokeratin system strictly dependent on the presence of desmosomes, which act as an organizing center for filament assembly. 106A-10a cells (also rich in desmosomes) and HeLa cells (showing instead a reduced number of desmosomes) have a cytokeratin system partially or totally independent from that of desmosomes, with different organizing centers.

Animals↗
PubMed1986

Marker evolution during the development of the rat mammary gland: stem cells identified by markers and the role of myoepithelial cells.

Using monoclonal antibodies and other immunological reagents we have identified characteristic markers for various epithelial cell types within the rat mammary gland. We have followed the evolution of cell types from the emergence of mammary ducts from the epidermis in the fetus to adulthood. Throughout mammary development some cells retain a group of markers which characterize the early stages of development. We have previously suggested that these cells are the stem cells for mammary development. In the adult, these cells are present in end buds and in the myoepithelial layer of ducts. We suggest that the myoepithelial layer, which we propose should be called the "basal" layer, contains several cell types, of which two are pluripotent. It contains the stem cells for mammary development, which also are present in end buds, and a precursor of ductules and alveoli. In the ducts, basal cells are probably also the precursor of luminal cells. We propose a scheme of mammary development.

Age Factors↗
PubMed1986

Distribution of developmental markers in rat mammary tumors induced by N-nitrosomethylurea.

We have examined the distribution of immunological markers in 55 intraductal carcinomas induced in rats by N-nitrosomethylurea using several different regimens of carcinogen treatment. The goal was to determine the possible relationship of the marker distribution to those existing at various stages of development of the mammary gland. We have also examined two passage lines in syngeneic rats derived from two of the tumors. The distribution of markers was not affected by the regimen of administration. The primary tumors were found to maintain the general topography of mammary ducts but with infoldings of the basal layer without accompanying stroma. We attribute this to an abnormal expression of the tendency of basal cells to migrate towards the lumen to generate luminal cells. The distribution of markers in the tumors was evaluated by identifying 13 special features of distribution that are common in tumors, using all-or-none criteria. The distribution of these features bears out the high heterogeneity of tumors in which the various features vary in a seemingly independent way. There is also heterogeneity within tumors, adjacent nodes often having different marker distributions. The distribution of the markers is related to that found in the early stages of mammary development. Because of this characteristic and of the fact that the tumors contain both basal and luminal cells, they must originate from multipotent cells, probably the stem cells present in end buds and ducts as already proposed by other work. As they develop, the tumors can both differentiate towards the adult type and dedifferentiate towards a fetal type. Of the two transplanted lines one retained the same general features of primary tumors in several passages, whereas the other evolved into a fusiform cell type with a marker distribution not seen at any stage of mammary development. Foci of similar cells were already present in the primary tumor, suggesting that the tendency to progress was already determined at an early stage. The fusiform cells are similar and probably equivalent to the fusiform cells that arise in vitro in cultures of rat mammary cancers.

Animals↗
PubMed1985

A monoclonal antibody to a mammary cell line recognizes two distinct subtypes of ovarian granulosa cells.

When screening supernatant fluids from hybridoma clones, Dulbecco and co-workers found that a mouse monoclonal antibody generated against a mammary tumor cell line showed rather striking high binding to rat oocytes and granulosa cells. In this study we have specifically investigated the reactivity of the monoclonal antibody (designated anti-OA-1) with granulosa cells during the differentiation process. This was accomplished using the two-step indirect immunocytochemical technique. When a primordial follicle is recruited to initiate growth, intense immunoreactivity appears in the surface membrane of the granulosa cells. As a follicle proceeds through the preantral stages, the plasma membrane of the granulosa cells is strongly positive for anti-OA-1 reactivity, and the granulosa appear as a homogeneous population. However, once Graafian follicle development is initiated, a major shift in anti-OA-1 immunoreactivity occurs among the granulosa cells. As the antrum expands, 75% of the granulosa in the mural region (those nearest the basal lamina) elongate and become negative for anti-OA-1. This is in contrast to the periantral and cumulus granulosa, which remain rounded and show strong anti-OA-1 reactivity up to the preovulatory stage. The disappearance of anti-OA-1 reactivity in the subpopulation of mural cells is specifically initiated by FSH and occurs very rapidly after a 12-h lag phase. After the loss of anti-OA-1 reactivity, the elongated mural granulosa cells express their terminal differentiated state by acquiring LH/hCG receptor, 3 beta-hydroxysteroid dehydrogenase activity and cytoplasmic lipid inclusions. By contrast, the periantral and cumulus granulosa, which remain positive for anti-OA-1, do not express these differentiated functions; however, they do differentiate ultrastructurally, indicating that they respond to the FSH signal. These results strongly suggest that a monoclonal antibody recognizes a major surface differentiation antigen in the granulosa cell. This antigen is under hormonal control and is inversely linked to expression of the terminal differentiation program in the granulosa cells. We anticipate that the monoclonal antibody will be a valuable probe to aide in the analysis of structure/function relationships in subpopulations of granulosa cells.

Animals↗
PubMed1985

Two monoclonal antibodies selective for human mammary carcinoma.

Mouse myeloma cells were fused with spleen cells from BALB/c mice immunized with the MCF-7 human mammary carcinoma cell line. Among hybridomas, two (3B18 and 15A8) were selected and cloned. Hybridoma 3B18 produces kappa-IgG1 antibodies that react with a cytoplasmic component of MCF-7 cells. In immunoperoxidase assays, 3B18 reacts with 27 of 31 specimens of human mammary carcinoma. It reacts most consistently with poorly differentiated and infiltrating ductal breast cancers, but it also reacts with isolated cells in 3 of 5 benign mammary pathological lesions with a variable distribution. The antibody does not react with normal mammary epithelium. It does not react with any normal human tissues, and it reacts with only one of 19 other cancers tested. Hybridoma 15A8 produces kappa-IgG1 antibodies that react with the surface membranes of the cells of two human breast cancer cell lines but not with a human fibroblast cell line. In immunoperoxidase assays, the antibody reacted with 28 out of 31 human mammary carcinomas. The antibody also reacts more weakly with normal human epithelial cells of breast, renal proximal tubule, skin, esophagus, and salivary gland, but no other normal tissue. The antibody was unreactive with 14 of 18 other malignant tissues tested. Since 3B18 and 15A8 detect antigens found predominantly in human mammary carcinomas and, possibly, distinguish overlapping categories of human mammary carcinomas, they may prove useful in determining the cellular lineage from which human mammary carcinomas arise, or they may have other clinical applications in breast cancer.

Antibodies, Monoclonal↗
PubMed1984

Microvillin: a 200-kilodalton protein in microvilli of rat mammary cells detected by a monoclonal antibody.

We have isolated a monoclonal antibody that was raised against rat mammary tumor cells. The antibody stains the lumenal edge of mammary duct epithelial cells. It immunoprecipitates a 200-kilodalton nonglycosylated, nonphosphorylated protein, which, by immunofluorescence and electron microscopy, can be localized in microvilli. In addition to microvilli of mammary ducts, this monoclonal antibody stains microvilli of the salivary ducts and of the renal nephron. It does not stain the microvilli of the intestinal brush border and shows a different tissue distribution than previously described proteins of this microvillus. We propose the name microvillin to distinguish the protein from other microvillar proteins. The presence of the protein distinguishes two classes of microvilli that are present in cells with possibly different transport functions.

Animals↗
PubMed1984

Lumen formation and redistribution of inframembranous proteins during differentiation of ducts in the rat mammary gland.

During the development of the rat mammary gland, ducts are formed from end-buds, which contain the stem cells. In this process a lumen is formed in the semisolid mass of the end-bud, and the cells acquire polarity. We have studied this process by following the localization of three inframembranous proteins present in the cells of both end-buds and ducts: microvillin, the microvillar protein p80, and the desmosomal plaque protein p205. We find that the development of ducts is accompanied by a redistribution of these proteins, which in immature parts of the end-buds are found together in the cell. Microvillin and p80 go together to the apical pole of the cells, in contact with the lumen, whereas p205 goes to the basal surface, in contact with cells of the myoepithelial lineage. The acquisition of polarity occurs at the same time as a lumen begins to form by local gaps between cells. It seems likely that the redistribution of the inframembraneous proteins is the consequence of the localization of surface glycoproteins that affect in opposite ways the adhesion between the cells.

Animals↗
PubMed1984

Developmental regulation of cytokeratins in cells of the rat mammary gland studied with monoclonal antibodies.

We have isolated two monoclonal antibodies to cytokeratins and determined their cell specificities. They display interesting localization within the rat mammary gland. One (1A10) shows specificity for myoepithelial cells; the other (24B42) is specific for lumenal cells at various stages of development. These two monoclonal antibodies and three others to cytokeratin previously isolated were used in conjunction with antibodies to myosin and collagen IV to confirm and extend our previous findings on epithelial cell types and development within the mammary gland.

Aging↗
PubMed1983

Epithelial cell types and their evolution in the rat mammary gland determined by immunological markers.

We have used immunological reagents for characterizing epithelial cell types in the adult rat mammary gland. Some were sera against purified proteins; others were monoclonal antibodies to mammary cells. On the basis of marker distribution we have identified 10 different cell types. We have established the developmental connection between these types by adopting the principle that cells displaying the same markers are directly related to each other. The results concur with those previously obtained by studying the growth response of mammary cells to the hormonal changes of the estrus cycle.

Animals↗
PubMed1983

Functional changes of intermediate filaments in fibroblastic cells revealed by a monoclonal antibody.

We describe reversible changes of intermediate filaments of fibroblastic cells associated with changes in the functional state of the cells. The changes are revealed by comparing the immunofluorescence patterns given by a monoclonal antibody and a polyclonal serum, both recognizing vimentin. The state of the filaments depends on culture density; this effect cannot be attributed to the nutritional state of the cells, their growth rate, or substances released into the medium. It seems to depend mainly on the aggregation of filaments during strong cell movements. The possible significance of these findings for the functional role of intermediate filaments is discussed.

Animals↗
PubMed1983

Tumor promoter binding to rat mammary cell cultures: role of receptors in the inhibition of dome formation.

We have studied the receptors for phorbol ester tumor promoters in a set of cultures of mammary cell lines derived from the same dimethylbenz(a)anthracene-induced tumor. These lines differ in cell morphology and in the ability to form domes either spontaneously or under the action of inducers. Tumor promoters inhibit dome formation. We asked whether receptors for tumor promoters could also mediate dome induction. We were unable to detect high-affinity receptors for the strong promoter, tetradecanoyl-12-phorbol-13-acetate (TPA) but could reveal receptors for the weaker promoter, phorbol-12, 13-dibutyrate (PDBU). Receptors were easily demonstrable during PDBU binding at 4 degrees; at 37 degrees, there was rapid disappearance of bound ligand, accompanied by its breakdown. Most cell lines have two classes of receptors with different affinities. Receptors for TPA could be demonstrated by competition with PDBU binding. In LA7 cells, prolonged exposure to PDBU caused down-regulation of the PDBU receptors with lower affinity. The half-effective doses of the dome-inhibiting effects are comparable to the KdS of the higher-affinity PDBU receptors measured by binding. TPA binds preferentially to the same class of receptors. This class of receptors, therefore, appears to be the one mostly involved in the anti-dome effect of either PDBU or TPA. Because dome inducers do not compete with PDBU binding, the PDBU receptors may not be responsible for dome induction.

Animals↗
PubMed1983

Genes in cancer.

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Animals↗