Biomedical subjects
Publications and source records attributed to P Monaghan.
Explore the source record for details and available documents.
The proportion of salmonella carriers among town-nesting herring gulls increased significantly from 2.1% in 1975-6 to 8.4% in 1979. The range of serotypes carried by herring gulls was similar to that causing infection in man, and it is likely that the gulls ingest these serotypes when feeding at untreated sewage outfalls on the coast. This is supported by the proportion of salmonella carriers being higher among first-year birds (9.7%) than among older birds (2.0%), as it is known that higher proportions of immature herring gulls feed on the coast. Herring gulls carrying salmonellas appeared healthy at the time of capture and at a later date it was assumed that they were not themselves infected. However, their habit of congregating in large numbers on reservoirs and rubbish tips and also at resting sites on farmland often far from feeding and roosting areas, multiplies the pollution problem and increases the potential health hazard for both man and farm stock. Herring gulls feed at a variety of sites and fly many miles from food source to food source and from feeding areas to the roost. Thus, even within the same day, there is the possibility of the transfer of salmonellas over a much wider area than previously considered.
The topographical distribution of type IV collagen, laminin, fibronectin, and the thymocyte differentiation antigen Thy-1 in the basement membrane of the lactating rat mammary gland was investigated. Small cubes of tissue, which had not been subjected to prior fixation or freezing, were incubated with monospecific or monoclonal antibodies to these proteins, and the antibodies were located by an indirect immunoperoxidase staining technique and observed in the electron microscope. The lamina densa stained uniformly with antibodies to type IV collagen and laminin. In addition, both laminin and type IV collagen were present in semiperiodic clusters that traversed the lamina lucida from the cell surface to the lamina densa. Fibronectin was present only in the semiperiodic clusters and not elsewhere in the basement membrane. These clusters were irregularly spaced along the cell surface and heterogeneous in size. It remains to be determined if these three proteins are present in the same clusters. Thy-1 was largely present on the lamina densa and not on the lamina lucida. The Thy-1 staining of the lamina densa occurred in discrete maxima and minima. These maxima occurred in regions adjacent to Thy-1-bearing stromal cells. Thus, the topographical distribution of proteins within a basement membrane varies in a nonrandom manner, and local factors can modify this distribution. We suggest that this topographical variability may play a role in cell recognition and signalling processes that occur across the basement membrane.
In the 100-day-old virgin and lactating rat mammary glands, type V collagen is mainly present in the interstitial connective tissue and in association with blood vessels. It is not present in the basement membrane region surrounding the ducts in mature virgin glands but is present in this region in neonatal and lactating glands. Ultrastructural localization of type V collagen reveals that it is mainly located on the basal surface (i.e., the surface in contact with the basement membrane) of epithelial but not myoepithelial cells. In addition, type V collagen is located on some interstitial collagen fibers and on a large number of granules that are in close proximity to the basal surface of both epithelial and myoepithelial cells. Immunofluorescence and biochemical studies indicate that several clonal mammary fibroblastic cell lines synthesize type V collagen in vitro. In some cell lines, type V collagen is secreted as an extensive fibrillar meshwork on the surface of the cells, whereas in other cell lines, it is secreted beneath the cells around their periphery. A number of mammary epithelial and myoepithelial-like cells, however, do not synthesize type V collagen in vitro.
The spontaneous production of elongated derivatives by cuboidal rat mammary epithelial cells was examined with the use of a series of single-cell clones grown in tissue culture. Four representative cell lines derived from a 7,12-dimethylbenz[a]anthracene-induced mammary tumor in an inbred WF rat were examined for morphologic stability, chromosome number, presence of immunoreactive fibronectin, laminin, prekeratin, and milk fat globule membrane (MFGM) antigens, ultrastructural characteristics, and tumorigenicity in syngeneic hosts. Conversion of cuboidal to elongated cells occurred by way of apparent morphologic intermediates, examples of which were isolated and cloned. Levels of immunoreactive fibronectin and laminin were greater in the elongated than the cuboidal clones, whereas the converse was true of prekeratin. MFGM antigens were present to a variable extent in all 4 clones. When grown on 0.3% collagen gels, cells of Rama 37 CL-A3 and Rama 37CS-A2 cuboidal clones exhibited surface microvilli and desmosomes. A minority of elongated cells contained microfilamental structures and pinocytotic vesicles similar to those seen in myoepithelial cells; the remainder lacked distinguishing ultrastructural features. After injection into syngeneic recipients, Rama 37 CL-A3 cuboidal line gave rise to glandular tumors consisting of cuboidal cells arranged in acinar structures, Rama 37 E5 elongated line induced spindle cell tumors, and Rama 37 CS-A2 and Rama 37 E8 lines induced tumors containing nests of mixed spindle and cuboidal cells. The majority of these tumors failed to metastasize.
A new human breast carcinoma cell line (PMC42) has been further characterized. The cells can grow either as monolayers or as floating cords of cells. The cords grow in suspension for long periods but may spontaneously attach and grow out to form a typical PMC42 monolayer. Ultrastructurally, the cells resemble breast ductal cells in many respects. Both epidermal growth factor (EGF) and prolactin induce ultrastructural changes, and lipid production is stimulated markedly by both factors. EGF also promoted the attachment of the floating cords and the growth of cells from these cords as monolayer cultures. The karyotype of the cord cells is different from that previously described for the monolayer cultures. Cord cells are hypodiploid (mode 39), whereas the monolayer cultures are subtriploid (mode 66). Although the ploidy is different, the karyotypes are related with 9 marker chromosomes being common to both populations. In addition, cultures in which cords have attached and in which cells are growing out as monolayers are bimodal with 10-20% of the cells becoming pseudotetraploid with a mode of 77.
Xenografts have been established in immune-suppressed mice from three embryonal cell carcinomas, two yolk sac carcinomas, and a 'seminoma-like' tumor that produces alphafetoprotein. The ultrastructural features of these xenografts have been studied in detail and compared with human germ cell tumors removed from patients. A spectrum of ultrastructural differentiation has been demonstrated for tumors of the same histologic classification (embryonal cell carcinoma) and within individual tumors. Cells with ultrastructural appearances intermediate between seminoma and yolk sac carcinoma and between embryonal cell and yolk sac carcinoma have been demonstrated, and their potential implications for tumor histogenesis are discussed.
A human cell line has been established from a transplantable xenografted human testicular tumor, which, both in the original tumor and in the xenograft, exhibited the histological characteristics of an undifferentiated malignant teratoma (embryonal cell carcinoma). The cells in culture were undifferentiated by biochemical, morphological, and ultrastructural criteria, growing as small islands of cells that tended to form aggregates at high density. The cells showed some variation in chromosome number with 30 to 40% of the cells having a normal human karyotype. The cells expressed high levels of alkaline phosphatase, which by heat inactivation and inhibition studies was 40 to 50% placental type alkaline phosphatase. None of the cultures produced human chorionic gonadotrophin, alphafetoprotein, carcinoembryonic antigen, or fibronectin, although at high cell densities plasminogen activator could be detected at low levels. Cell surface studies showed that the cells shared antigens with the murine embryonal carcinoma cell line F9, expressed beta 2-microglobulin at very low and variable levels, and bound the lectin peanut agglutinin. These studies suggest that this cell line has some of the characteristics described for murine embryonal carcinoma cell lines.
With the use of immunofluorescent and immunocytochemical techniques on histologic sections of mammary glands from inbred WF rats, the thymocyte differentiation antigen (Thy-1) was identified partially on and immediately adjacent to the myoepithelial cells of ducts and alveoli. This antigen was absent from epithelial cells lining such structures. Some fibroblastic cells external to these structures also bore Thy-1. During glandular development the intensity of fluorescence due to the binding of Thy-1 antibodies increased as the myoepithelial cells matured. Similarly, mammary adenocarcinomas induced by 7,12-dimethylbenz[a]anthracene (DMBA) and N-methyl-N-nitrosourea contained elongated, presumptive myoepithelial cells that demonstrated varying degrees of Thy-1 immunofluorescence. This phenomenon was qualitatively mimicked by cultured cell lines from normal and DMBA-induced tumors. Elongated myoepithelial-like and fibroblast-like cells possessed Thy-1, whereas the cuboidal epithelial cell lines failed to express this antigen on the cell surface. Measurement of the relative number of Thy-1 molecules per cell by antiserum absorption techniques suggested that a neoplastic stem cell line, rat mammary (Rama) 25, contained about 3 X 10(5) Thy-1 molecules per cell in a form not directly accessible at the cell surface to anti-Thy-1 antibodies; the number of cryptic Thy-1 molecules was reduced when this cell line differentiated to alveolar-like cells in culture. However, when this cell line differentiated to myoepithelial-like cells, approximately 5 X 10(6) molecules per cell were exposed to anti-Thy-1 antibodies with a concomitant reduction of the cryptic pool. Morphologic maturation of elongated, myoepithelial-like cell line variants was also accompanied by increased surface Thy-1 fluorescence. Thus some of the myoepithelial cells in normal glands and tumors may arise by differentiation of epithelial stem cells and a spectrum of maturation states of the myoepithelial cell may exist as monitored by cellular morphology and surface Thy-1 expression.
Ten cell lines of human squamous carcinomas of the tongue and larynx have been established from surgical specimens removed from 36 unselected patients, in order to provide systems for investigating the invasive and tissue-destructive capacity of squamous carcinomas of the head and neck. The morphology, ultrastructure and growth characteristics of the 10 lines are described. Detailed cytogenetic analysis of the first 4 lines indicates that each is karyotypically unique, with no evidence of cross-contamination. Nine of the 10 cell lines secrete immunoreactive beta human chorionic gonadotrophin (beta-hCG) in the culture medium. No correlation was demonstrated between the ability of the cell lines to secrete plasminogen activator and their capacity to grow in soft agar or as xenografts in immune-deficient mice.
Five tumour cell lines have been derived from a primary squamous carcinoma of the tongue, from 2 subsequent local recurrences, and from 2 lymph-node metastases--all from the same patient. While the cell lines shared many morphological and biochemical characteristics, those derived from recurrences and metastases appeared to be less differentiated, were less well organized in culture, and displayed fewer desmosomes and tonofilaments than cells in the primary tumour line. A recurrent line showing greatest morphological divergence from the primary tumour line also demonstrated the greatest differences at the ultrastructural level, in increased production of plasminogen activator and in the composition of cell-surface glycoproteins.
A clonal, myoepithelial-like cell line has been obtained from a primary culture established from the mammary gland of a 7-d-old rat. In a number of respects, this cell line, termed Rama 401, resembles the myoepithelial cells of the mammary gland, especially when grown on floating collagen gels. The cells grow as multilayers on the gel surface and form branching structures that do not appear to contain a lumen. They are rather elongated, with irregular-shaped, flattened nuclei that contain large amounts of peripheral chromatin. Elongated processes project from the cell surface and numerous membrane pinocytotic vesicles can be seen. The cytoplasm is filled with linear arrays of 5- to 7-nm filaments with occasional dense foci. Cell junctions with associated 8- to 11-nm tonofilaments are also observed. Immunofluorescence techniques reveal actin and myosin filaments and also intermediate filaments of both prekeratin and vimentin types. Rama 401 cells secrete an amorphous material that, when an immunoperoxidase technique is used, stains with antibodies to basement membrane-specific type IV collagen. Localized densities of the cell membrane, which resemble hemidesmosomes, are located adjacent to these extracellular deposits. Immunofluorescence staining and immunoprecipitation techniques reveal that the cells also synthesize two other basement membrane proteins, laminin and fibronectin. The type IV collagen consists of two chains with molecular weights of 195,000 and 185,000.
A xenograft line, HX 53, has been established in immune-suppressed mice from a specimen of a lymph node metastasis in a patient with a histological diagnosis of seminoma but with markedly raised circulating levels of alpha-fetoprotein. Histological, immunocytochemical, and ultrastructural studies of this xenograft line have suggested that a solid variant of yolk sac carcinoma may exist, which morphologically resembles seminoma, or that a continuum of differentiation exists between seminoma and yolk sac carcinoma.
N-Methyl-N-nitrosourea (MNU) given iv to rats 50-55 days old induced mammary tumors in 70% of F344/N and 91% W/ICRF inbred females with mean latency periods of 149 and 93 days, respectively. Reduction of the MNU dose did not affect tumor incidence in W/ICRF rats. Of the mammary tumors, 98% were classified histologically as adenocarcinomas, which grew progressively. Primary tumors of nonmammary origin were detected at low incidence. Upon histologic examination, no evidence was found for metastases of either the mammary or other primary tumors. No evidence for tumor-induced hypercalcemia was found. Oophorectomy at the time of MNU administration prevented tumor development; oophorectomy when at least 1 tumor/animal was palpable caused growth delay or regression. All MNU-induced and 7,12-dimethylbenz[a]anthracene-induced mammary tumors tested contained cytoplasmic estrogen receptor (ER) at similar concentrations and were indistinguishable histologically. MNU-induced tumors in F344 rats were transplantable and retained ER through three transplantations.
The routine preparation of gram quantities of lobuloalveolar and ductal structures from human reduction mammaplasties by treatment with collagenase is described. When cultured on plastic or glass surfaces these structures give rise, initially, to epithelial sheets consisting of cells which retain many morphological characteristics of myoepithelial cells and do not stain with an antiserum which reacts with the surfaces of the lining epithelium of breast ducts and lobulo-alveoli. Subsequently, cells which resemble the lining epithelial cells migrate from these structures and react strongly with the antiserum. Cell proliferation in these cultures is minimal. Explants of major ducts dissected from mastectomies produce vigorously proliferating epithelial sheets which contain morphologically similar cells not identifiable as either lining epithelium or myoepithelium, although nearly all the cells in direct contact with the medium react with the lining epithelium specific antiserum.
The preparation of highly purified monolayer cultures of human cytotrophoblast cells essentially free of stromal and syncytial cells is described. Such cells subsequently form multinucleate syncytial cells in vitro. This is accompanied by the synthesis of heat-stable alkaline phosphatase and beta-HCG. A significant proportion of the multinucleate cells that form in monolayer cultures of early placentae arise as a result of an amitosis. It is suggested that this in vitro behavior may reflect an in vivo process.
Mice immunologically deprived by thymectomy, cytosine arabinoside treatment and whole-body irradiation were used to study the growth of human tumours as xenografts. 10/16 melanoma biopsies, 4/13 ovarian carcinoma biopsies and 3/6 uterine cancer biopsies grew as serially transpllantable xenograft lines. The tumour lines were studied through serial passages by histology, histochemistry, electron microscopy, chromosome analysis, immune fluorescence, growth rate measurement and mitotic counts. They retained the characteristics of the tumours of origin, with the exception of loss of pigmentation in two melanomas, histological dedifferentiation in the uterine carcinomas, and increased mitotic frequency and growth rate in some melanomas. It was concluded that this type of animal preparation is as useful as alternative methods of immunological deprivation, or as athymic nude mice, for the growth of human tumour xenografts, at least for some experimental purposes.
The tissue culture characteristics of a primordial cyst are described and compared with normal oral mucosa. The potential application of this method to other aspects of oral biology is suggested.