Interactions of normal, dysplastic, and malignant mammary epithelial cells with fibronectin in vivo and in vitro.

Previous studies have demonstrated that normal and malignant mouse mammary cells are indistinguishable in many surface-related properties that often denote transformation of other cell types such as fibroblasts. In the present investigation, the interactions of normal, dysplastic, and malignant mammary epithelial cells with fibronectin in tissues and cultures were examined by indirect immunofluorescence. Cells lining the lumina of ducts and alveoli in normal and dysplastic mouse and human mammary tissues abutted a layer of fibronectin along their basal surfaces that included the region of the basement membrane and the underlying stroma. Moreover, double staining for keratin and fibronectin revealed that myoepithelial cells were surrounded by the matrix protein. In contrast, tumor cells in adenocarcinomas and ductal carcinomas were not directly associated with fibronectin. The accumulation of fibronectin in primary cultures prepared from mouse mammary tissues paralleled the distribution seen in vivo. A matrix of fibronectin formed beneath normal and preneoplastic mammary cells within 4 to 6 days after plating, whereas tumor cells were negative, regardless of the age or density of the culture. This correlation with in vivo results did not extend to cells of established mammary tumor culture lines which readily accumulated pericellular networks of fibronectin. Addition of exogenous fibronectin to primary cultures enhanced formation of a basal matrix by normal cells but had no effect on the negative status of the tumor cells. The results indicate that mammary tumor cells in tissues and in primary cultures have an altered capacity to interact with fibronectin. However, this alteration is not necessarily expressed by established mammary tumor cell lines.

Glutathione transferase activity and isoenzyme composition in primary human breast cancers.

The human glutathione transferases (GSTs) are a multigene family of detoxication enzymes with patterns of expression that are both tissue specific and genetically determined. Changes in the levels of one or more GST isoenzymes have been associated with the development of anticancer drug resistance in cultured cell lines. In this study, total GST activity and GST isoenzyme composition have been determined for 45 primary human breast carcinomas using a 1-chloro-2,4-dinitrobenzene substrate assay and Western blotting, respectively. The GST activity ranged from 5-208 mU/mg protein with a mean of 67 mU/mg protein (+/- 44 SD). GST-pi) isoenzyme protein was detectable on Western blots in 44 of 45 samples. Mu Class GST protein was detected in 18 of 38 samples and undetectable in 20 of the 38 samples tested. By polymerase chain reaction analysis of genomic DNA, the absence of mu class GST in breast tumors was determined to be due to the deletion of the gene for GST-mu in the DNA of those tumors. None of the 43 primary human breast cancer samples tested contained detectable alpha class GST protein. Neither the total GST activity of tumor samples, the quantity of GST-pi protein, nor the presence or absence of mu class GST correlated with other factors known to be of prognostic significance including tumor size, nodal status, estrogen receptor protein positivity, or progesterone receptor protein positivity. Substantial differences exist among primary breast carcinomas in both the amount of GST activity and GST isoenzyme composition. However, these are not tightly linked either to tumor stage or to hormone receptor status. Whether the levels of these enzymes are independent predictors of either risk of recurrence or response to anticancer therapy has yet to be tested directly.

In vitro assay for human breast cancer hormone responsiveness. A study of 25 cases with clinical correlation.

PIP: The in vitro effects of cortisol, estradiol, progesterone, and testosterone on the uptake of tritiated thymidine into the tumor cells of 25 women with primary or metastatic breast cancer were studied and compared with the objective clinical response to ablative endocrine surgery. A correlation between the clinical results with endocrine therapy and the in vitro assay results obtained in 23 of the 25 patients. 8 patients who did not respond to clinical endocrine therapy did not respond in vitro. A significant clinical response (p less than .05) to ablative endocrine surgery, lasting for at least 6 months, was observed in 9 of 15 patients whose tumors responded in vitro. Precise determination of the predictive capacity of the assay will require a blind prospective evaluation.^ieng

Identification of mouse mammary epithelial cells by immunofluorescence with rabbit and guinea pig antikeratin antisera.

Few markers are available to identify the three types of mammary epithelial cells--ductal, alveolar, and myoepithelial--especially in pathological conditions and in cell cultures. We have used antisera to human keratins in immunofluorescence to facilitate the identification of the three mouse mammary epithelial cell types. In frozen tissue sections and primary cell cultures, a rabbit antikeratin antiserum specifically stained cytoplasmic filaments in all three types of epithelial cells. A guinea pig antiserum against the same keratin preparation, however, reacted preferentially with filaments in myoepithelial cells and readily detected this cell type in normal, dysplastic, and malignant mammary tissues and cell cultures. Neither antisera reacted with fibroblasts or any other mesenchymal cells. The combined use of the two antikeratin antisera thereby permits rapid surveys of tissue sections and cultures for the localization of not only all epithelial cells but also the subpopulation of myoepithelial cells. Moreover, when mammary cultures established from late-pregnant or lactating mice were stained simultaneously with guinea pig antikeratin and rabbit anticasein antisera, three populations of epithelial cells were mutually exclusive: those stained by anticasein antiserum, those stained by guinea pig antikeratin antiserum, and those stained by neither, consistent with properties of alveolar, myoepithelial, and ductal cells, respectively. These antisera thus offer a tool for studying different epithelial cell types during mammary development, tumorigenesis, and malignant progression.

Vector potential of houseflies (Musca domestica) for Helicobacter pylori.

The mode of transmission of Helicobacter pylori is unknown. Since viable bacteria have been shown to be excreted in feces from infected individuals and houseflies habitually develop and feed on excrement, we hypothesized that flies ingest and harbor H. pylori and, in turn, contaminate the human environment. This study examined the possible vector potential of houseflies (Musca domestica) for H. pylori. Caged houseflies were exposed to freshly grown H. pylori on agar plates. After a 6-h feeding period, the plates were removed and were replaced with sterile petri dishes containing a droplet of sterile brucella broth. At regular intervals, small numbers of houseflies were removed for microbiological and histological analysis, and the petri dishes were replaced with fresh sterile plates with fresh drops of brucella broth. The flies' bodies, the flies' dissected alimentary tracts, and excreta on the petri dishes were cultured for H. pylori, whose identity was confirmed by the urease, catalase, and oxidase reactions and Gram staining. In contrast to control flies, viable H. pylori could be isolated from external surfaces for up to 12 h and from gut and excreta for as long as 30 h after the initial feeding period. After 30 h other gram-negative bacteria overgrew the cultures of samples from all locations tested, rendering the selective culture of H. pylori colonies impossible. Histological analysis revealed Helicobacter-like organisms in the gut lumen and attached to intestinal epithelial cells. We conclude that houseflies can harbor viable H. pylori on their bodies and in their intestinal tracts. They are also able to disseminate viable H. pylori in excreta, and they may therefore present a significant reservoir and be a vector in the transmission of H. pylori.

Structural distinctions among human breast epithelial cells revealed by the monclonal antikeratin antibodies AE1 and AE3.

Two monoclonal antikeratin antibodies, AE1 and AE3, were used in indirect immunocytochemistry to examine keratin expression in normal, benign proliferative, and malignant human breast epithelium. Both antibodies reacted strongly with most luminal cells in ducts and acini of normal gland. While AE1 did not stain myoepithelium, AE3 recognized myoepithelial cells of ducts but not acini, implying a cytoskeletal difference between the myoepithelium of these two components. Moreover, the antibodies reacted differently with the myoepithelium of intracanalicular as compared with pericanalicular types of fibroadenomas. Tumour cells of infiltrating ductal carcinomas with a prominent intraductal component stained more homogeneously with AE1 and AE3 than those without intraductal growth. The results provide evidence for two phenotypes of myoepithelial cells and for the presence of cryptic keratin epitopes in human breast epithelial cells. The finding that neither AE1 nor AE3 is a universal detector of these cells has important clinical and experimental implications.

Histologic and mammographic specificity of risk factors for benign breast disease.

This study evaluates the effects of potential risk factors for benign breast disease (BBD) with special attention to the histologic and mammographic specificity of the effects. Cases were 172 women with BBD that underwent biopsy; controls were 134 women free of breast signs or symptoms. All cases and controls had undergone mammography. For all types of BBD combined, parity, use of oral contraceptives, and use of exogenous estrogen after menopause were strongly protective, whereas obesity and early menarche were weakly protective. Family history of breast cancer was virtually unrelated to BBD. The protective effect of parity was stronger for BBD with intralobular or extralobular fibrosis, and with mammographic homogeneous density or large nodular densities, than it was for BBD without these characteristics. Similar relations with the histologic and mammographic features were observed for obesity. These findings suggest that some risk factors for BBD have effects that are related to specific features of its morphology.

Relationship between mammographic and histologic features of breast tissue in women with benign biopsies.

Mammograms and histologic slides of a group of 320 women who had breast symptoms and a biopsy without cancer being found were reviewed. The mammographic features assessed were the parenchymal pattern and extent of nodular and homogeneous densities. In addition to the pathologic diagnosis, the histologic features assessed included epithelial hyperplasia and atypia, intralobular fibrosis, and extralobular fibrosis. Among premenopausal women, those with marked intralobular fibrosis were more likely to have large (3+ mm) nodular densities on the mammogram. Among postmenopausal women, epithelial hyperplasia or atypia was related to having nodular densities in at least 40% of the breast volume. In both groups, marked extralobular fibrosis was related to the presence of homogeneous density on the mammogram. We conclude that mammographic nodular densities may be an expression of lobular characteristics, whereas homogeneous density may reflect extralobular connective tissue changes.