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Reproducible subculture of adult human prostatic epithelial cells from normal, benign hyperplastic and malignant tissue has been achieved. Cholera toxin is the key component in the culture system, but use of an optimal basal medium (PFMR-4) supplemented with a high level of serum in collagen-coated dishes also improves growth and serial propagation. [ABSTRACT FROM AUTHOR]

Normal mouse mammary epithelial cells in primary culture can be passaged as viable single cells using 0.5 to 1.0 mg/ml pepsin in Hanks' salt solution. After 5 min the pepsin treatment preferentially removes fibroblasts, leaving a monolayer of purified epithelial cells that can be removed by pipetting and transferred to new culture vessels or injected into animals. [ABSTRACT FROM AUTHOR]

A number of liver epithelial cell cultures were established from 10 to 12-d-old sucklings, 6 to 8-wk-old young adults, or from 2 to 18-month-old partially hepatectomized rats. Improvements in the methods for cell isolation and culture yielded replicating cells from every experiment and they were maintained for different periods with regular passages. The proliferative potential in vitro of adult rat liver cells could be increased if the rats were subjected to partial hepatectomy before cell isolation. In the early passages, the majority of the cells were found to have a true diploid karyotype as studied by the Giemsa-banding technique. Under the culture conditions described, a very high percentage of cells remained in the diploid range for, in most cases, at least 4 months and in some cases for up to 6 months. Afterward, the karyotype was unstable, but no 'crisis' period was seen before the cells became aneuploid. Until this time, the growth characteristics of the cells also followed a normal pattern showing density dependent inhibition of division and a lack of markers associated with malignancy. The cultured liver cells exhibited a number of liver specific properties when they were maintained as a confluent monolayer. The early passages of diploid epithelial cell cultures derived from normal and regenerating rat liver are good models for studies of the regulation of cell division and the changes that are related to carcinogenesis. [ABSTRACT FROM AUTHOR]

Defined culture conditions for routine clonal growth of normal human adult bronchial epithelial cells have been developed. Serum and feeder cell requirements were abrogated by: (a) optimizing the calcium concentration in nutrient medium, MCDB 151; (b) supplementing with purified factors (epidermal growth factor, 5 ng/ml; insulin, 5 µg/ml; transferrin, 10 µg/ml; hydrocortisone, phosphoethanolamine and ethanolamine, each at 5×10 M; and trace elements); and (c) coating the surface of the culture dish with a mixture of fibronectin, collagen, and bovine serum albumin. Endothelial cell growth supplement (100 µg/ml) and retinoic acid (3 × 10 M) further enhanced growth, whereas cholera toxin was nonmitogenic and serum supplementation (>2%) markedly reduced the growth rate. Using the defined system, dissociated cultures of bronchial epithelial cells, obtained from more than 15 donors, have been subcultured at clonal densities with a colony forming efficiency of 3 to 4%. In addition, high density cultures have been subcultured more than five times with four to six population doublings per passage. The features of this system permit pathobiologic investigations of bronchial epithelial cells, e.g., aging, differentiation, and carcinogenesis using conditions that isolate the results from the influence of serum, feeder cells, and other undefined factors. [ABSTRACT FROM AUTHOR]

A new cell line, SS78, was established from a primary renal cell carcinoma of a Caucasian male. The tissue was dispersed with collagenase, and viable cells were separated by flotation on a Ficoll-Hypaque gradient. In culture, the SS78 cells retained a distinct epithelial morphology, and no fibroblastlike cells were seen. The cultured cells were aneuploid with a modal chromosome number of 80 and had several marker chromosomes. Inoculation of the cultured cells into athymic nude mice caused tumors at the sites of inoculation. [ABSTRACT FROM AUTHOR]

Immunofluorescence and immunoperoxidase methods were used to identify constituents and products of cultured human bronchial epithelial cells and fibroblasts. Epithelial cells, but not fibroblasts, from patients with blood Types A or B reacted to the respective antisera to either the A or B blood group antigens. However, neither the epithelial cells nor the fibroblasts from patients with blood type O[H] reacted with the anti-H antisera. Epithelial cells in primary culture reacted with antibody to prekeratin proteins from human stratum corneum and fibroblests did not react. Moreover, keratin filaments were assembled in vitro from proteins isolated from the epithelial cells. These immunological and biochemical data support previous morphological observations that human bronchial epithelial cells in primary culture shift progressively from a mucociliary epithelium to a keratinizing epithelium. Epithelial cells and fibroblasts could also be identified by their reactivity to anticollagen antibodies. Fibroblasts reacted strongly with antibodies to Types I and III collagens and epithelial cells did not. On the other hand, epithelial cells reacted weakly with antibodies to Type IV collagen and fibroblasts were completely negative. Both epithelial cells and fibrobalsts reacted with antibody to fibronectin; however, the distribution of fibronectin differed in the two cell types. In epithelial cells, fibronectin was restricted to the cell surface, whereas in fibroblasts it was found on the cell surface and in the extracellular matrix where fibrils of fibronectin were both cell associated and deposited on the surface of the dish where the fibroblasts had migrated. [ABSTRACT FROM AUTHOR]

The individual effects of seven hormones on the in vitro growth rate of different classifications of human mammary epithelium were compared. Hormones used were: 17β-estradiol, estriol, progesterone, hydrocortisone, testosterone, prolactin, and growth hormone. Cell cultures included three established breast cell lines and primary monolayer cultures established form breast fluids and excised mammary tissue from 40 women and 4 men. Specimens comprised three classifications: normal, nonmalignant atypical, and malignant. Growth was quantitated in situ and expressed as population doubling time. Principal findings were: (a) estrogens, prolactin, and growth hormone stimulated growth of normal cells more frequently than growth of malignant cells, whereas testosterone and hydrocortisone stimulated growth of malignant cells more frequently than growth of normal cells; (b) cells cultured from nonmalignant atypias generally showed hormone response profiles intermediate between those of normal and malignant cells; (c) progesterone stimulated the growth of cells from malignant specimens but not the growth of cells from normal and nonmalignant atypical samples. [ABSTRACT FROM AUTHOR]

Explants of adult human bronchus were cultured in CMRL-1066 medium supplemented with heat-inactivated fetal bovine serum, hormones, antibiotics, and either 0.0, 0.1 1, or 10 m M putrescine. The outgrowth of bronchial epithelial cells was stimulated in medium containing 1 m M putrescine, a concentration that partially inhibited the outgrowth of fibroblasts. In medium containing 10 m M putrescine, the outgrowth of epithelial cells was similar to that observed in the control medium, but the outgrowth of fibroblasts was completely inhibited for periods of at least 4 weeks. When 10 m M putrescine was added to cultures of bronchial fibroblasts, the fibroblasts were not killed. These results suggest that human bronchial epithelial cells have a higher requirement for putrescine for growth than fibroblasts, and the molecular basis for this observation is under investigation. [ABSTRACT FROM AUTHOR]

We have tested the sensitivity of a cloned rat hepatocyte line, RL-PR-C, to aflatoxin B and benzo(a)pyrene as a function of population-doubling level. The cells were much more sensitive to the cytotoxic action of these agents subsequent to 230 population doublings. This sensitivity corresponded to the enhanced inducibility of arylhydrocarbon hydroxylase activity by 3-methylcholanthrene. [ABSTRACT FROM AUTHOR]

Chronic exposure of a cloned rat hepatocyte culture (RL-PR-C) to a subtoxic, sublethal dose of aflatoxin B resulted in malignant transformation. Continuous exposure to aflatoxin B caused increasing tumorigenic potential as tested by back injection into isogenic animals. Control cultures exhibited spontaneous transformation, although approximately 20 passages beyond the chemically induced event. Neither aflatoxin-treated nor control cultures exhibited cytopathological morphology, formation of cell foci, growth in soft agar, or irregular fibroblast-like growth patterns that could be specifically related to the onset of tumorigenic potential. In general, those parameters commonly used to monitor fibroblast cultures for transformation in vitro were not applicable for assessing the tumorigenic potential of these epithelial cells. Karyotypic analyses revealed no specific chromosomal aberrations associated with aflatoxin treatment; however, chromosomal instability was a property of the tumorigenic cell populations. Injection of both aflatoxin-treated and control cultures at passage 56 resulted in tumors indicative of both carcinoma and sarcoma indicating to us the multipotency of these epithelial cells transformed in vitro. [ABSTRACT FROM AUTHOR]

An operational criterion for the identification and isolation of epithelial-like (E) cells, based on their ability to cover and protect, a collagen gel from the action of collagenase, has been developed. The E cells isolated by this collagenase-separation technique (CST) exhibited the ultrastructural features, including desmosomes and abundant tonofilaments, that are considered characteristic of this cell type. Unlike confluent cultures of fibroblast-like (F) cells, E cells were not found to have large external transformation-sensitive (LETS) protein on their surface membranes. The CST provides a nondestructive, and efficient means of identifying and isolating E cells from mixed populations. [ABSTRACT FROM AUTHOR]

A cell line and subline with epithelial characteristics were established from mouse mammary hyperplastic alveolar nodules (HAN). The cells do not grow in suspension cultures in vitro or form tumors in vivo. The cells do produce significant amounts of C-type and A-type virus and low amounts of plasminogen activator. [ABSTRACT FROM AUTHOR]