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L B Chen

Publications and source records attributed to L B Chen.

At least 163 records · Page 9Linked to original sources
PubMed1982

Role of small t antigen in the acute transforming activity of SV40.

A plasmid, pHR402, containing SV40 sequences that include a truncated early region bearing an intact t-coding sequence and a functionally intact late region, was introduced into thymidine kinase deficient (tk-) mouse L cells by cotransformation with a cloned tk gene. tk+ cotransformants synthesized SV40 t but not T antigen, and no truncated T-coding sequence products were detected. The viral sequences of pHR402 were reconstituted as a virus in COS1 cells, and acute infection of untransformed mouse cells with this viral stock (SV402) also led to the appearance of t but not T or a truncated T. Abortive transformation assays of such infected cells were negative, as were those performed on the same cells infected with either of two viral mutants (dl883 and dl884), each of which leads to T but not t synthesis. However, mixed infection with SV402 and either dl883 or dl884 led to a clear abortive and permanent transformation response. Thus, at least in part, t and T appear to function in a complementary fashion in eliciting transformation expression by SV40-infected cells.

Animals↗
PubMed1982

Decreased uptake and retention of rhodamine 123 by mitochondria in feline sarcoma virus-transformed mink cells.

A reduce uptake and retention of the mitochondria-specific membrane potential probe rhodamine 123 by feline sarcoma virus (FeSV)-transformed mink fibroblasts (64F3) has been detected. The decreased accumulation of rhodamine 123 by 64F3 mitochondria is not due to abnormal plasma membrane dye permeability, since after microinjection of the dye these cells are still unable to retain the dye at levels comparable to the untransformed parental cells, CCL 64. Nigericin, an ionophore that mediates an electrically neutral exchange of protons for potassium ions resulting the elimination of the pH gradient across the mitochondrial membrane and a compensatory increase in mitochondrial membrane potential with continued respiration, increases both the dye uptake and the retention time in transformed 64F3 cells. These results suggest that mitochondria in FeSV-transformed mink cells may have an abnormally low mitochondrial membrane potential accompanied by a relatively high pH gradient. Since anioic metabolites such as pyruvate and glutamate are accumulated by mitochondria in proportion to the delta pH across the mitochondrial membrane, the abnormal mitochondria described here may contribute to the abnormal metabolic state of FeSV-transformed cells.

Animals↗
PubMed1982

Unusual retention of rhodamine 123 by mitochondria in muscle and carcinoma cells.

Mitochondria in cardiac muscle cells and myoblast-fused myotubes display unusually long (3-5 days) retention times of rhodamine 123, a mitochondria-specific fluorescent probe, in living cells. Among 50 keratin-positive carcinoma or transformed epithelial cell lines tested, mitochondria with prolonged rhodamine 123 retention are detected in most of the transitional cell carcinoma, adenocarcinoma, and chemical carcinogen-transformed epithelial cell lines and in some squamous cell carcinoma lines but not in any oat cell carcinoma lines. The presence of mitochondria having unusual dye retention may be useful for diagnosis and exploitable for chemotherapy of certain human carcinomas.

Animals↗
PubMed1982

Cell types associated with fibronectin in long-term mouse bone marrow cultures.

The formation of fibronectin matrix was studied in long-term mouse bone marrow cultures. Stromal and hematopoietic cells were observed in situ under phase contrast optics and quantified according to their staining characteristics on smear preparations. Surface fibronectin was demonstrated by indirect immunofluorescence. While only stromal and no hematopoietic cells participated, various stromal cell types differed in their expression of cell surface fibronectin: Reticulum cells contributed the major portion of fibronectin matrix. Elongated, meshwork-forming histiocytes expressed some surface fibronectin, while the flattened, macrophagic histiocytes remained fibronectin negative. These findings were recapitulated during regeneration of scrape wounds in the adherent layers. Isolated fibronectin matrix did not support hematopoietic cell adherence or maintenance, although it had marked effects on stromal cells.

Adipose Tissue↗
PubMed1982

Localization of a Mr 52,000 keratin in basal epithelial cells of the mouse bladder and expression throughout neoplastic progression.

In this study, we report the isolation of a Mr 52,000 keratin from a nontumorigenic bladder epithelial cell line and its localization, by immunofluorescence, in bladder frozen sections at different stages of neoplastic progression and in various carcinogen-transformed bladder epithelial cells and human bladder carcinoma-derived cell lines. The results showed that: (a) the Mr 52,000 keratin is present in basal epithelial cells of the normal bladder but absent from the intermediate and superficial epithelial cell layers; (b) hyperplastic bladder lesions, induced in mice after the administration of butyl-(4-hydroxybutyl)-nitrosamine, resulted primarily from the proliferation of cells in the basal compartment; (c) butyl-(4-hydroxybutyl)nitrosamine-induced bladder carcinomas displayed differential staining with the anti-Mr 52,000 keratin antiserum reflecting divergent differentiated status within a tumor and a subpopulation of cells with reduced overall keratin expression; (d) primary cultures of bladder carcinomas revealed a subpopulation of cells with limited filamentous keratin as was observed in vivo; (e) mouse bladder epithelial cell lines transformed in culture by a chemical carcinogen showed a loss or reduction in expression of the Mr 52,000 keratin; (f) two human bladder carcinoma cell lines displayed very limited expression of the Mr 52,000 keratin. Although the loss or reduction in expression of a specific keratin is unlikely to be responsible for transformation, it may contribute to the heterogeneity in the differentiated state and morphology of bladder epithelial cells during neoplastic progression both in vivo and in culture.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1982

Induction of cytoskeleton-associated proteins during differentiation of human myeloid leukemic cell lines.

Alterations in the expression of proteins associated with the cytoskeletal framework during differentiation of two human myeloid leukemia cell lines were analyzed by two-dimensional gel electrophoresis of Triton-insoluble cellular framework fractions. During in vitro differentiation of HL60 (human promyelocytic leukemia line) and U937 (human monocytoid leukemia line), several new cytoskeleton-associated (CSK) proteins are induced. All of these CSK proteins are also present in freshly isolated normal granulocytes and macrophages. One of these differentiation-induced proteins comigrates with vimentin. There are several differentiation-sensitive proteins, i.e., those that are no longer synthesized upon differentiation. The changes in CSK protein synthesis during differentiation of HL60 and U937 cells do not seem to be related to drug treatment per se since exposure to conditioned medium from phytohemagglutinin-stimulated lymphocytes as well as to dimethyl sulfoxide and 12-O-tetradecanoylphorbol-13-acetate results in the production of many similar proteins. In vitro conditions that do not result in differentiation of HL60 and U937, such as cultivation in serum-free medium, do not induce the CSK proteins that we describe. A notable finding in this study is that all of the qualitative changes in the proteins synthesized during differentiation are detected in the cytoskeletal (Triton-insoluble) fraction, whereas only minor quantitative alterations are observed in the Triton-insoluble extract. The changes in CSK protein components occur in an orderly fashion. Vimentin, an intermediate-filament protein, is synthesized in large amounts prior to changes in cellular morphology and the induction or loss of other CSK proteins. Vimentin may play an important role in the reorganization of the cytoskeleton to support the process of differentiation. The other CSK proteins are synthesized sequentially along with the morphological and functional changes during differentiation. These model systems, therefore, present an opportunity to investigate the role of specific cytoskeletal components in cellular differentiation.

Carcinoma, Small Cell↗
PubMed1981

Detection of phosphotyrosine-containing 34,000-dalton protein in the framework of cells transformed with Rous sarcoma virus.

Phosphotyrosine-containing 34,000-dalton protein is detected by treatment of a two-dimensional gel of cellular framework with 1 M NaOH at 40 degrees C for 1 hr. The alkali-resistant 32PO4-labeled 34,000-dalton protein is detected in various cell lines transformed by Rous sarcoma virus but not in lines transformed by simian virus 40, polyoma virus, herpes simplex II virus, adenovirus type 2, or chemical carcinogens. In addition, interferons or fibronectin matrices have no detectable effect on the phosphorylation of the 34,000-dalton protein in Rous sarcoma virus-transformed cells.

Animals↗
PubMed1981

Monitoring of relative mitochondrial membrane potential in living cells by fluorescence microscopy.

Permeant cationic fluorescent probes are shown to be selectively accumulated by the mitochondria of living cells. Mitochondria-specific interaction of such molecules is apparently dependent on the high trans-membrane potential (inside negative) maintained by functional mitochondria. Dissipation of the mitochondrial trans-membrane and potential by ionophores or inhibitors of electron transport eliminates the selective mitochondrial association of these compounds. The application of such potential-dependent probes in conjunction with fluorescence microscopy allows the monitoring of mitochondrial membrane potential in individual living cells. Marked elevations in mitochondria-associated probe fluorescence have been observed in cells engaged in active movement. This approach to the analysis of mitochondrial membrane potential should be of value in future investigations of the control of energy metabolism and energy requirements of specific biological functions at the cellular level.

Animals↗
PubMed1981

Expression of keratin and vimentin intermediate filaments in rabbit bladder epithelial cells at different stages of benzo[a]pyrene-induced neoplastic progression.

Rabbit bladder epithelium, grown on collagen gels and exposed to the chemical carcinogen benzo[a]pyrene, produced nontumorigenic altered foci as well as tumorigenic epithelial cell lines during 120-180 d in culture. Immunofluorescence studies revealed extensive keratin filaments in both primary epithelial cells and benzo[a]pyrene-induced altered epithelial foci but showed no detectable vimentin filaments. The absence of vimentin expression in these cells was confirmed by two-dimensional gel electrophoresis. In contrast, immunofluorescence staining of the cloned benzo[a]pyrene-transformed rabbit bladder epithelial cell line, RBC-1, revealed a reduction in filamentous keratin concomitant with the expression of vimentin filaments. The epithelial nature of this cell line was established by the observation that cells injected into nude mice formed well-differentiated adenocarcinomas. Frozen sections of such tumors showed strong staining with antikeratins antibodies, but no detectable staining with antivimentin antibodies. These results demonstrated a differential expression of intermediate filament type in cells at different stages of neoplastic progression and in cells maintained in different growth environments. It is apparent that the expression of intermediate filaments throughout neoplastic progression is best studied by use of an in vivo model system in parallel with culture studies.

Animals↗
PubMed1981

Association of phenotypic reversion of transformed cells induced by interferon with morphological and biochemical changes in the cytoskeleton.

The distribution of microfilament bundles, intermediate filaments, microtubules, vinculin plaques, and extracellular fibronectin matrices was examined by indirect immunofluorescence in X-ray-transformed C3H/10T1/2 cells. The transformed phenotype correlated with a lack of actin cables and vinculin-containing plaques. Prolonged treatment of these transformed cells with interferon resulted in a reversion of the transformed phenotype which was associated with the reappearance of extensive microfilament bundles together with vinculin-containing plaques. Both interferon-treated and untreated transformed cells displayed a similar distribution of mitochondria, intermediate filaments, microtubules, and fibronectin. Significant qualitative and quantitative differences between the two types of cells were found in the proteins associated with the cytoskeletal framework by two-dimensional gel analysis.

Animals↗
PubMed1980

Specific disruption of vimentin filament organization in monkey kidney CV-1 cells by diphtheria toxin, exotoxin A, and cycloheximide.

We have examined the effect of diphtheria toxin, Pseudomonas aeruginosa exotoxin A, and cycloheximide on the CV-1 cell cytoskeleton. Within a few hours after producing an inhibition of cellular protein synthesis, all these agents specifically disrupted the organization of the vimentin filament system with no discernable effect on microtubules or microfilaments during the period of observation. Furthermore, just as the inhibition of protein synthesis by cycloheximide is reversible, so was the disruption of vimentin filaments by cycloheximide.

Animals↗
PubMed1980

Localization of mitochondria in living cells with rhodamine 123.

The laser dye rhodamine 123 is shown to be a specific probe for the localization of mitochondria in living cells. By virtue of its selectivity for mitochondria and its fluorescent properties, the detectability of mitochondria stained with rhodamine 123 is significantly improved over that provided by conventional light microscopic techniques. With the use of rhodamine 123, it is possible to detect alterations in mitochondrial distribution following transformation by Rous sarcoma virus and changes in the shape and organization of mitochondria induced by colchicine treatment.

Animals↗
PubMed1980

Studies of fibronectin matrices in living cells with fluoresceinated gelatin.

We have used fluorescein isosthiocyanate-conjugated gelatin (FITC-gelatin) (1 mg/ml) to localize cell surface fibronectin in unfixed live cells in cultures. FITC-gelatin stains the fibronectin matrix on primary cultures of rat and chick embryo fibroblasts as well as untransformed, established cell lines. In live cultured cells, fibronectin in many areas of the extracellular matrix is inaccessible to antibody and cannot be visualized by immunofluorescence staining. In contrast, fibronectin in these areas is fully stainable by FITC-gelatin. At a low concentration (20 micrograms/ml), FITC-gelatin stains the fibronectin matrix of primary cultured cells but not of "untransformed" established cell lines. SEM can detect only the matrix stainable with the low concentration of FITC-gelatin, such as that expressed by primary chick embryo fibroblasts. The binding of fibronectin to the extracellular matrix is very stable and FITC-gelatin remained bound to the matrix for at least 10 d in culture. Radioiodinated gelatin has been used to quantitate the level of cell surface fibronectin in living normal and transformed cells. FITC-gelatin appears to be a useful probe for studying the fibronectin of living cells in culture.

Animals↗
PubMed1979

Malignant behaviour of three adenovirus-2-transformed brain cell lines and their methyl cellulose-selected sub-clones.

Three adenovirus-2-transformed rat embryo brain cell lines and their methylcellulose-selected sub-clones were examined for fibronectin expression, anchorage-independent growth, saturation density, T antigen expression and morphology. Tumorigenicity studies were carried out on newborn and ATS immunosuppressed syngeneic rats and congenitally athymic nude mice. With one exception the methylcellulose sub-clones contained significantly fewer fibronectin-positive cells than the parent lines; a number of sub-clones contained no fibronectin-positive cells. Methylcellulose selection did not always alter cell morphology, saturation density or anchorage-independent growth as compared with parent lines. However, the methylcellulose sub-clones were considerably more malignant than the parent cell lines as measured by invasion and metastasis in nude mice. No in vitro characteristic correlated with malignant behaviour.

Adenoviridae↗