Adeline J. Hackett

Growth of diploid cells from breast cancers

Cell cultures were derived from normal and cancerous breast tissues and from metastases by methods that selected for relatively adherent epithelial aggregates. Karyotypic analyses of first or second passage cultures yielded predominantly normal diploid cells. Nonclonal aberrations were more common in tumor-derived than in normal cultures. Three of the cultures that originated from metastases were characterized by abnormal clones. These results support observations based on DNA content, which indicate that a considerable fraction of breast cancers are composed predominantly of diploid cells. They differ greatly from chromosomal findings in long-term cultures of tumor effusions and thus emphasize the karyotypic diversity that can be found in tumors from a single tissue of origin--the breast.

A possible morphologic basis for the autointerference phenomenon in vesicular stomatitis virus

Abstract A physical particle which could account for the phenomenon of autointerference in vesicular stomatitis virus (VSV) is described. This particle is present in virus prepared by the multicyclic growth method, following inoculation with a high multiplicity of virus. Architectural details of the New Jersey serotype of VSV are presented.

The biology of breast cancer at the cellular level

Two properties seem fundamental to cancer; heterogeneity and progression (Foulds (1975) Academic Press, New York; Heppner et al. (1979) Commentaries on Research in Breast Disease, Vol. 1 (Bulbrook, R. and Taylor, D.J., eds.), pp. 177-191, Plenum Press, New York). Relatively little is understood about the premalignant stages of human breast disease in vivo. When the disease manifests as invasive carcinoma, its behavior exhibits great diversity, sometimes metastasizing rapidly, while in other cases 10-30 years pass before metastases proliferate. Here we review various aspects of breast cancer in vivo and consider how they predict properties of breast cancer found in culture. All of the experiments are consistent with the hypothesis proposed by Nowell (1976) Science 194, 23-28, that a fundamental aspect of malignancy is an increased genetic instability and that many of the cells within tumors are nonviable results of genetic instability. We suggest that most of the viable cells within primary breast carcinomas are diploid and are not yet capable of aspects of metastatic spread. What these cells have attained is an increased propensity for genetic instability which enables them to generate randomly aneuploid but frequently lethal genetic configurations. Occasionally one of these altered genomes is associated with the ability to proliferate at a metastatic site. This hypothesis implies that metastases from various patients may have arisen by divergent pathways and may also be divergent in many other aspects of their physiology, unrelated to malignancy. Such extreme heterogeneity may hamper attempts to understand fundamental aspects of malignancy. Hence we suggest that the less anaplastic and less divergent diploid cells within the primary carcinomas might be an important resource to gain insights into the critical alterations that are responsible for initiating frankly malignant behavior.

Immunolocalization of a human basal epithelium specific keratin in benign and malignant breast disease

SummaryThis report describes the immunocytochemical localization of a human basal- or myoepithelial-specific antikeratin antibody in benign and malignant breast disease. Reactivity patterns with this antibody have demonstrated the lack of myoepithelial or basal epithelial participation in most benign breast specimens examined including those displaying cystic disease, fibrosis, or hyperplasia. However, in specimens of sclerosing adenosis, strong reactivity with the majority of cells in most ducts suggests a major participation of the myoepithelial cell type. Analysis of 118 breast carcinoma specimens has demonstrated strong, homogeneous reactivity in 4% of the specimens, suggesting a role for the basal epithelial cell in malignancy of the human mammary gland and implications for the prognosis of such tumors. Antigenic characterization of the malignant and benign mammary specimens which are uniformly reactive with the antibody has demonstrated the presence of a 51 kd keratin polypeptide not found in the non-reactive specimens.

Biological properties of human melanoma cells in culture

SummaryThree human melanoma cell lines derived from one primary and two metastatic tumors from three different patients were characterized for growth properties usually associated with malignant transformation; these include cell morphology, growth rate, saturation density, growth in semisolid media, colony-forming ability on contact-inhibited monolayers of normal fibroblasts and epithelial cells, and tumorigenicity in immunosuppressed mice. Variations in expression of aberrant properties were evident among the lines. One of the metastatic lines satisfied all the parameters of malignancy tested and the other showed a number of these properties, whereas the primary essentially fulfilled only one. These results suggest that cultured melanoma cells reflect the clinical variability often observed among melanoma patients and the metastatic melanoma seems to display a higher degree of malignant transformation than the primary.

The separation of infectious and autointerfering particles in vesicular stomatitis virus preparations

Abstract Vesicular stomatitis virus (VSV) infections produce two sizes of particles that differ not only in length, but also in infectivity and autointerference properties. The two forms have been separated by sucrose density gradient centrifugation after partial purification by differential centrifugation, enzyme, and fluorocarbon treatment. Peaks of radioactivity of 32 P-labeled preparations coincided with the two light-scattering bands. Infectivity was associated with the lower band which contained long (L) rods. Autointerference was associated with the upper band which contained short (S) rods.

Differential retention of rhodamine 123 by breast carcinoma and normal human mammary tissue.

SummaryWe have qualitatively evaluated the retention of the fluorescent dye rhodamine 123 by malignant or non-malignant breast epithelial cells in passively-infused fresh surgical specimens. Our findings demonstrate a microscopically-visible increase in the ability of primary and metastatic tumor cells to retain the dye, as compared to non-malignant epithelium. Some variability in fluorescence intensity was seen within and between tumor specimens. The optimal length of incubation in the presence of the dye was critical in achieving differential fluorescence intensity between normal and malignant cells. This method of examining rhodamine 123 uptake and retention in tissue explants provides a reliable means for direct, comparative visualizationin situ of any tissue and its associated disorders. The results of this study also demonstrate the validity of extending the use of liophilic, cationic compounds such as rhodamine 123 as antitumor agents, from model systems to the treatment of malignant disease.

Biological characteristics of two plaque variants of vesicular exanthema of swine virus, type E54

Abstract Two lines of vesicular exanthema virus have been isolated which differ from each other in the size of the plaque produced. Several of the in vitro biological characteristics of these lines have been examined, to account for the difference in size. The minute variant was adsorbed more rapidly than the large variant to swine kidney cells and also multiplied more rapidly within these cells. Ninety per cent of the virus produced was cell associated. Average yields per cell were in excess of 1000 plaque-forming units with both variants grown at pH 8. In multicyclic growth the concentration of bicarbonate affected the amount of virus produced by infected cells, and the pH of the medium affected the release of virus after maturation. At pH 7 the release of the minute variant was small and extracellular virus was only 1–10% of that which remained in or attached to the cells as long as 20 hours after the peak of virus growth. Release of the large-plaque variant from the cells was more rapid and more complete both at acid and alkaline pH levels. Stability of the large variant at 36° was greater at pH 7 and 8. The differences in rate and efficiency of release of virus from infected cells and in pH stability are believed to account, in part, for the observed difference in plaque morphology.

Early expression of vimenti in human mammary cultures

SummaryThe intermediate filaments of most epithelial cells in vivo consist solely of cytokeratins. Using monoclonal antibodies to vimentin or keratin, we have examined the expression of vimentin in homologous specimens of frozen tissue sections and primary cultures of normal human mammary epithelium. In frozen sections, only epithelial cells reacted with the antikeratin antibody, whereas antivimentin reactivity was associated with stromal cells. All epithelial cultures were positive for cytokeratin and in addition coexpressed vimentin as strongly as cultured fibroblasts and as early as the 4th d after initiation of the culture. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis of cytoskeletal preparations of secondary cultures of normal mammary epithelium have also demonstrated the appearance of a moiety identical to the vimentin found in cultured fibroblasts. Our observations are consistent with the hypothesis that vimentin expression is induced, possibly as a result of changes in cell shape or growth rate, when cells are freed from three-dimensional restirctions imposed by the tissue of origin

Isopycnic zonal centrifugation and characterization of the mouse mammary tumor virus (MTV) in different gradient solutions

Abstract The buoyant density of MTV in several different gradient solutions was determined, using a single pool of MTV-infected milk as the source of virus. Increased ultraviolet absorbance and detectable MTV B-particle antigenicity correlated with the positions of discrete light-scattering bands formed during centrifugation. MTV isodensities were found to be similar to those reported for other RNA tumor viruses. Centrifugation of MTV-positive preparations in preformed gradients containing sucrose or potassium tartrate resulted in the formation of two distinct light-scattering bands. Electron microscopic examination showed that only those bands at the higher buoyant density contained characteristic MTV particles. The bands of lower isodensity consisted of viruslike particles having considerable variability in size and shape, which may represent incomplete MTV particles. The properties of isopycnically centrifuged MTV-free milk samples were clearly distinguishable from those of MTV-positive samples; no discrete light-scattering bands were formed, and no viruslike particles were observed in electron micrographs.