Expression of GPC3, an X-linked recessive overgrowth gene, is silenced in malignant mesothelioma.

Gene expression changes in rat asbestos-induced malignant mesothelioma (MM) cells were investigated by differential mRNA display. A mRNA transcript identified by this approach was abundant in normal rat mesothelial cells but not expressed in rat MM cell lines. Northern blot analysis confirmed that this transcript is uniformly silenced in rat MM cell lines and primary tumors.

Fibronectin expression and organization in mesothelial and mesothelioma cells.

Mesothelial cells are believed to be the progenitor cells for malignant mesothelioma, a tumor associated with exposure to asbestos and other mineral fibers. Little is known regarding fibronectin (Fn) function in mesothelial and mesothelioma cells. Fn RNA, protein levels, and localization were assessed in secondary cultures and later passages of spontaneously immortalized rat pleural mesothelial (NRM) cells and in neoplastic cell lines derived from asbestos-induced mesotheliomas.

Mucin production by SPOC1 cells--an immortalized rat tracheal epithelial cell line.

An airway epithelial mucous goblet cell line would be useful towards understanding mechanisms underlying the common problem of respiratory mucus hypersecretion. SPOC1 is a novel rat tracheal epithelial (RTE) cell line that developed cytologic features suggestive of mucous goblet cells when grown in tracheal grafts in vivo (Am. J.

Phenotype and differentiation potential of a novel rat tracheal epithelial cell line.

In this report we described the establishment and characterization of a continuous rat tracheal epithelial (RTE) cell line spontaneously derived from secondary RTE cell cultures. Designated SPOC1, this cell line is nontumorigenic and maintains a diploid karyotype with specific, nonrandom chromosomal alterations involving chromosomes 1, 3, and 6. SPOC1 cells demonstrate decreased requirements for peptide growth factors, compared with primary RTE cells.

Autocrine growth stimulation by transforming growth factor alpha in asbestos-transformed rat mesothelial cells.

Although the association between asbestos exposure and mesothelioma development has been established for decades, very little is known regarding the molecular mechanism(s) by which asbestos fibers induce this disease. In this series of experiments, the potential for transforming growth factor alpha (TGF-alpha) to act as an autocrine growth factor in transformed mesothelial cells was examined in rats, a model system frequently used to assess the tumorigenic potential of fibrous particulates. Both asbestos-transformed cells and spontaneously transformed cells expressed functional EGF receptors, although only the asbestos-transformed cells expressed TGF-alpha.

Alterations in the localization of F-actin, fibronectin, and thrombospondin occur prior to neoplastic transformation in rat tracheal epithelial cells.

Structural glycoproteins and cytoskeletal proteins play a major role in the regulation of cellular organization and function. Changes in the structure and function of these proteins are involved in the cascade of events which lead to neoplastic transformation. We evaluated RNA levels, protein localization, and organization of selected proteins in an in vitro model system of respiratory carcinogenesis to examine alterations in cell architecture.

Regulation of normal and transformed tracheobronchial epithelial cell proliferation by autocrine growth factors.

Several peptide growth factors have been identified as autocrine regulators of normal tracheobronchial epithelial cells, including the transforming growth factors alpha and beta. Using in vitro tissue culture models of multistep neoplastic transformation as well as cell lines derived from human bronchogenic tumors, a significant number of growth factors have been found to function as autocrine factors for neoplastic tracheobronchial cells as well. In several instances, the regulation of the autocrine pathway is disrupted in neoplastic tracheobroncial cells compared with their normal counterparts.

Cellular uptake of phosphorothioate oligodeoxynucleotides is negatively affected by cell density in a transformed rat tracheal epithelial cell line: implication for antisense approaches.

Antisense oligodeoxynucleotides have been used to specifically inhibit the expression of a variety of genes. In the process of testing phosphorothioate oligodeoxynucleotides (S-oligos) as specific inhibitors of transforming growth factor alpha synthesis, we observed that the efficiency of cellular uptake of S-oligos was not constant throughout the culture period but decreased as the cell densities increased. Our data suggest that the mechanism by which oligonucleotides enter cells is negatively affected by increased cell density.

Epidermal growth factor dependence and TGF alpha autocrine growth regulation in primary rat tracheal epithelial cells.

We have examined dependence of primary rat tracheal epithelial (RTE) on exogenous epidermal growth factor (EGF) and determined whether a TGF alpha autocrine pathway is operating in these cells. Primary RTE cells plated in serum free media (SFM) without EGF and bovine pituitary factor (BPE) show little proliferation compared to cultures propagated in media containing EGF/BPE (CSFM). Removal of EGF/BPE shortly after plating, however, results in significant proliferation, although plateau cell densities are reduced and cell morphology is significantly altered compared to cells propagated in CSFM.

Alterations in growth factor pathways in multistage carcinogenesis of rat tracheal epithelial cells.

The role of peptide growth factors in the process of multistage carcinogenesis of rat tracheal epithelial (RTE) cells was assessed by examining growth factor requirements and expression of growth factors and their receptors in normal and transformed RTE cells. Transformed RTE cell lines show decreased requirements for bovine pituitary extract, insulin and epidermal growth factor compared to primary RTE cells in culture. An autocrine role for TGF alpha in transformed RTE cells is suggested by data showing TGF alpha production and decreased proliferation in the presence of TGF alpha antisera and TGF alpha/EGF receptor kinase inhibitor.

Role of TGF alpha and its receptor in proliferation of immortalized rat tracheal epithelial cells: studies with tyrphostin and TGF alpha antisera.

We have shown in the present study and in studies reported previously that preneoplastic and neoplastic rat tracheal epithelial (RTE) cell lines express TGF alpha and do so regardless of the mechanism by which they were transformed. In order to determine whether TGF alpha is an autocrine growth regulator of immortalized RTE cells, we have examined the function of TGF alpha/EGF receptors and the growth requirements for TGF alpha in these cells. The level of immunoprecipitated TGF alpha/EGF receptor protein in immortalized RTE cells was similar to or less than levels in primary RTE cells, indicating that chemically induced transformation of RTE cells does not involve overexpression of TGF alpha/EGF receptors.

Abnormalities of growth factor systems in transformed airway epithelial cells.

The role of the peptide growth factors transforming growth factor alpha (TGF alpha) and transforming growth factor beta (TGF beta) in the regulation of proliferation of normal and transformed airway epithelial cells was studied. Normal as well as transformed rat tracheal epithelial (RTE) cell cultures secrete similar amounts of TGF alpha during logarithmic growth. Once normal RTE cell cultures reach the plateau phase of growth, they down-regulate TGF alpha expression at the RNA and protein level; in contrast, transformed cells do not down-regulate TGF alpha.

Abnormalities in growth regulation of transformed rat tracheal epithelial cells.

We have examined the changes in growth regulation which take place during multistage transformation of rat tracheal epithelial (RTE) cells exposed to chemical carcinogens in vitro. Transformed RTE cells demonstrate: (1) altered negative growth factor sensitivity and (2) altered positive growth factor dependence and gene expression compared with primary RTE cells in culture. Primary RTE cells are sensitive to the growth inhibitory effects of retinoic acid and transforming growth factor beta (TGF beta).

The effect of azelastine and some other antiasthmatic and antiallergic drugs on calmodulin and protein kinase C.

The antiallergic and antiasthmatic drug, azelastine, interacts strongly with calmodulin (but not bovine serum albumin) as determined by an indirect assay; it also moderately inhibited the Ca2+-calmodulin-dependent enzyme bovine brain phosphodiesterase. Ketotifen was less active than azelastine in both assays of calmodulin reactivity and both drugs were less active than the recognized calmodulin inhibitor, W-7. Neither azelastine nor ketotifen had any inhibitory effect on the Ca2+- and phospholipid-dependent protein kinase C.

Protein kinase C inhibition by plant flavonoids. Kinetic mechanisms and structure-activity relationships.

Protein kinase C (PKC) from rat brain was inhibited by plant flavonoids in a concentration-dependent manner depending on flavonoid structure. Of the fifteen flavonoids studied, fisetin, quercetin and luteolin were the most potent, while hesperetin, taxifolin and rutin were among the least potent. The flavonol fisetin was almost 100% inhibitory at a concentration of 100 microM.

Altered growth factor dependence and transforming growth factor gene expression in transformed rat tracheal epithelial cells.

The role of peptide growth factors in neoplastic progression of transformed rat tracheal epithelial (RTE) cells was assessed by examining growth factor requirements and expression of growth factor and growth factor receptor genes in normal and transformed RTE cells. Neoplastically transformed cell lines showed decreased requirements for bovine pituitary extract, insulin, and epidermal growth factor compared to normal primary RTE cells. Neoplastic RTE cell lines expressed significantly increased levels of transforming growth factor alpha (TGF alpha) RNA and secreted TGF alpha into the media, suggesting an autocrine role for this growth factor.

Thiazide diuretics inhibit chloride absorption by rabbit distal colon.

In order to investigate the cellular mechanisms of action of thiazide diuretics, the effect of diuretic and nondiuretic thiazide compounds on Cl- absorption across rabbit distal colon was assessed in tissues mounted in Ussing chambers. This epithelium absorbs Cl- via an active electroneutral transport process. Net 36Cl- absorption across short-circuited tissues was decreased 53, 36 and 20% after addition of 10(-4) M trichlormethiazide, bendroflumethiazide or hydrochlorothiazide, respectively, to the mucosal bathing solution.

Localization of chloride secretion in rabbit colon: inhibition by anthracene-9-carboxylic acid.

The substituted aromatic compound anthracene-9-carboxylic acid (A-9-C) was used to inhibit active Cl- secretion by the epithelium of short-circuited rabbit distal colon. Tissues were mounted in Ussing chambers and stimulated to secrete Cl- by the addition of 1 mM dibutyryl adenosine 3',5'-cyclic monophosphate to the serosal bath. Results of 36Cl-flux measurements showed that the addition of 0.