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

We have isolated a mouse monoclonal antibody that, upon immunohistochemical localization in frozen sections, displays specificity for human myoepithelial cells in the resting mammary gland, sweat glands, and salivary glands. Furthermore, this antibody was strongly and homogeneously reactive with frozen sections of 3 of 60 breast carcinoma specimens. Using immunolocalization techniques in conjunction with polyacrylamide gel electrophoresis, we have determined that the reactivity of this monoclonal antibody is directed toward a 51,000-dalton keratin polypeptide. The potential uses of this antibody in the prognosis of human mammary carcinoma and in understanding the role of the myoepithelium in development and differentiation are discussed.Myoepithelial cells are derived from ectoderm and are known to exhibit both epithelial and mesenchymal characteristics (1, 2). They are situated between acinar or ductal luminal cells and the basal lamina in a number of secretory glands such as breast, lacrimal, eccrine, and apocrine sweat glands and various salivary glands (3).We report here on a monoclonal antibody directed toward a cytoskeletal keratin that is unique to the myoepithelial or "basal" layer of epithelial cells in human mammary ducts, sweat glands, and salivary glands. Using polyacrylamide gel electrophoresis and immunoblotting, we have found the apparent molecular weight of this basal epithelium-specific keratin to be 51,000 daltons. The antibody is generally nonreactive with luminal epithelium, stroma, muscle, fat, and vasculature. Therefore, it appears to be a useful tool in studying the role of the myoepithelium in the normal development of the breast, during lactation and involution, and in malignancy and metastasis.Recently, considerable advances have been made in tumor diagnosis by immunocytochemical typing of intermediate filaments. For example, carcinomas are characterized by antibodies to cytokeratins; sarcomas of muscle cells, by anti-desmin; nonmuscle sarcomas, by anti-vimentin; and gliomas, by anti-glial fibrillary acidic protein (for review, see ref. 4). The myoepithelium-specific monoclonal antibody described in this report has allowed us to define and characterize subclasses of human mammary carcinomas that would not be possible to distinguish by conventional techniques. The potential application of this antibody in the prognosis of human mammary carcinoma is discussed. MATERIALS AND METHODSImmunization. BALB/c mice raised under pathogen-free conditions were injected via subcutaneous or intravenous routes for 5 weeks at weekly intervals with whole-cell preparations of a secondary culture of an infiltrating ductal breast carcinoma. For each inoculation, 106 cells, which had been cultured as previously described (5), were prepared by scraping the monolayer with a rubber policeman into phosphate-buffered saline. Splenocytes from immunized animals were fused with SP2/0 mouse myeloma cells in the presence of polyethylene glycol (Mr 4000; Merck) by the procedure of Kohler and Milstein (6). Monocl...

Section: Discussionmentioning

confidence: 99%

Monoclonal antibody that defines human myoepithelium.

Dairkee¹,

Blayney²,

Smith³

et al. 1985

“…As primary antibodies, we used the mouse monoclonal antibody lA10 (basal cytokeratin; ref. 10), 24B42…”

Section: Methodsmentioning

confidence: 99%

“…10), and 91316 (microvillin; ref. 11) always in conjunction with a polyclonal rabbit serum specific either for laminin (courtesy of E. Engvall, La Jolla Cancer Research Foundation) or for myosin (courtesy of S. J.…”

mentioning

confidence: 99%

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

Dulbecco¹,

Armstrong²

1988

Self Cite

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.The genetic mechanisms by which cancer initiates are known (1), but those that drive the progression of cancer through a series of stages to full malignancy (2), mainly through local invasion and metastasis, are unknown. The conditions allowing cells to metastasize have been studied extensively (for a review, see ref.3) by taking advantage of experimental systems based on cell cultures. In contrast, for lack of a suitable model system, the study of invasion has been limited to basic histological observations.An experimental system widely used as a model for human breast cancer is mammary cancer induced in rats by Nmethyl-N-nitrosourea (4). However, these tumors are not regularly invasive (5) or metastatic (6). This difference between rat and human tumors might be due to the vast difference in developmental time, from a few months in the rat to decades in humans. To increase the time available for the expression ofthe invasive potential, we have transplanted the tumors serially to isogeneic rats, an approach already used for detecting the oncogenic potential of mouse hyperplastic alveolar nodules induced in the mammary gland by the mouse mammary tumor virus (7). The observations made show the usefulness of the experimental system, clarify several aspects of progression and invasion, and establish a similarity between the rat tumors and human mammary cancers. We show that tumors are heterogeneous in their ability to invade neighboring tissues, which behaves like a stochastic process of varying probability. This result is in accord with the finding that heterogeneity is the hallmark of tumors, whether one considers the presence of specific products (5) or the ability of premalignant lesions to develop into neoplasias (8). MATERIALS AND METHODSFifteen Wistar/Furst rats received a subcutaneous injection of 5 mg of N-methyl-N-nitrosourea per 100 g of body weight at the age of 5 weeks and a second identical injection 4 weeks later. The animals were maintained with standard die...

“…The presence of E cadherin in the domes probably indicates that it allows the formation of junctions of a special nature, enabling the cell layer to be lifted by the underlying hydrostatic pressure produced by the pumping function. The expression of a cytokeratin is also a sign of a differentiation process, because cytokeratins of various types are expressed in the cells of mammary ducts at various stages of differentiation (31). Cytokeratin 8, which is expressed in the domes, is frequently associated with tubular structures in other organs (32).…”

mentioning

confidence: 99%

The rat gene homologous to the human gene 9–27 is involved in the development of the mammary gland

Zucchi

Montagna

Susani

et al. 1998

We have developed a model system for studying differentiation in the mammary gland, by using two clonal cultures deriving from a rat breast adenocarcinoma. They differ in the ability to form domes, structures the significance of which is unknown. By using the subtractive cDNA library approach, we isolated a cDNA that is highly expressed in the dome-forming cells, and identical to the rat8 gene and highly homologous to the human 9-27 gene. Antisense treatment of the dome-forming cells specifically and reproducibly abolishes dome formation, while forced expression of the gene in non-dome-forming cells causes morphological changes suggestive of ''f lat'' domes. In situ hybridization on rat tissues shows that the gene is expressed in epithelia, especially in those forming tubular structures, suggesting a relatedness between these structures and domes. Cytokeratin 8 and E cadherin are strongly expressed in the domes but not outside them, suggesting that the rat8 gene triggers the cells to express molecules that tighten the lateral connections between the cells; the process is likely to parallel that occurring during the differentiation of the mammary gland.