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Biomedical subjects

A E Levine

Publications and source records attributed to A E Levine.

At least 19 recordsLinked to original sources
PubMed2001

OrCGDB: a database of genes involved in oral cancer.

The Oral Cancer Gene Database (OrCGDB; http://www.tumor-gene. org/Oral/oral.html) was developed to provide the biomedical community with easy access to the latest information on the genes involved in oral cancer. The information is stored in a relational database and accessed through a WWW interface. The OrCGDB is organized by gene name, which is linked to information describing properties of the gene. This information is stored as a collection of findings ('facts') that are entered by the database curator in a semi-structured format from information in primary publications using a WWW interface. These facts include causes of oncogenic activation, chromosomal localization of the gene, mutations associated with the gene, the biochemical identity and activity of the gene product, synonyms for the gene name and a variety of clinical information. Each fact is associated with a MEDLINE citation. The user can search the OrCGDB by gene name or by entering a textword. The OrCGDB is part of a larger WWW-based tumor gene database and represents a new approach to catalog and display the research literature.

Databases, Factual↗
PubMed2000

Digital reviews in molecular biology: approaches to structured digital publication.

MOTIVATION: It is widely appreciated that it is no longer possible for biomedical research scientists to keep up with as much of what is published in their field as they ought. One solution to this problem is to increase the efficiency of information use by moving away from the classical browsing model for scientific information dissemination towards an information on demand model which would allow researchers to access information quickly and efficiently only as they need it. The most common approach to this goal has been to use information retrieval technology to improve access to text databases of biomedical information. We are interested in exploring an alternative; encoding this information for storage in structured databases for efficient retrieval. RESULTS: Two small databases described here are test beds for development of structured digital publication software; the Tumor Gene Database, containing information about genes which are the sites for cancer-causing mutations, and the Mammary Transgene Database, containing information about expression of transgenes in agriculturally important animals. Both have been successfully searched by users and edited by curators via the World Wide Web.

Algorithms↗
PubMed1999

Increased transforming growth factor-alpha levels in human colon carcinoma cell lines over-expressing protein kinase C.

Transforming growth factor-alpha (TGF-alpha) is synthesized as a membrane-bound precursor protein, pro-TGF-alpha, that is converted to a soluble form by 2 endoproteolytic cleavages. Several factors have been implicated in the regulation of the second rate-limiting step, including protein kinase C (PKC). Earlier results indicated a potential role for the conventional class of PKC isozymes in the observed increase in TGF-alpha in the conditioned media of 2 human colon carcinoma cell lines. The present study addresses the potential role of specific PKC isozymes in this process using sense and anti-sense expression vectors for PKC isozymes. Two human colon carcinoma cell lines, HCT 116 and GEO, were transfected with plasmids, leading to the over-expression of PKC-alpha, -betaI or -betaII; and the secretion of TGF-alpha into the conditioned medium was determined. Over-expression of either PKC-betaI or PKC-betaII in these cell lines enhanced the levels of TGF-alpha in the media 2- to 5-fold. Over-expression of PKC-alpha did not alter the amount of TGF-alpha in the media to a significant extent. Transfection of HCT 116 cells with the anti-sense PKC-betaI cDNA resulted in a reduction in PKC-betaI protein expression. This was accompanied by a decrease in the amount of TGF-alpha in the conditioned media. Our results indicate that modulation of PKC-beta protein levels alters the amount of TGF-alpha found in the conditioned media from these colon carcinoma cells.

Colonic Neoplasms↗
PubMed1997

Transforming growth factor-beta response and expression in junctional and oral gingival epithelial cells.

The junctional (JE) and oral gingival (OGE) epithelium show distinct morphological phenotypes and express different cell surface and keratin markers. Transforming growth factor-beta (TGF-beta) has been shown to stimulate extracellular matrix formation and inhibit proteolytic matrix degradation in periodontal wound healing. To elucidate potential roles of TGF-beta in gingival epithelial regeneration and reattachment, the present study examined the effects of TGF-beta on JE and OGE cell growth and determined the patterns of expression of mRNAs for the TGF-beta isotypes beta 1, beta 2 and beta 3 and TGF-beta receptor types I, II and III. Primary cell cultures were initiated from JE and OGE and the cell phenotypes confirmed using monoclonal antibodies to specific keratins. TGF-beta induced a significant growth inhibition in OGE cells derived from 6 different patients with a mean inhibition of 46% and a range of 16-70% (p = 0.031). Although responses varied between patients, in general maximum inhibition occurred at 10 ng/ml TGF-beta. JE cells from 5 patients showed no significant growth inhibition by TGF-beta (p = 0.125). Greater expression of TGF-beta 2 and receptor type I mRNA was found in OGE than JE cells and thus appeared to be associated with differentiating epithelial cells. JE cells expressed more TGF-beta type II receptor specific mRNA than did OGE cells, but TGF-beta 1 mRNA expression was similar in JE and OGE cells. JE or OGE cultures derived from 2 of 3 patients showed expression of mRNA for the TGF-beta type III receptor. TGF-beta 3 mRNA was not detected in any of the JE or OGE samples examined. The greater sensitivity of OGE than JE to the growth inhibiting effects of TGF-beta correlated with higher expression of receptor type I mRNA which, together with the type II receptor, is required for sensitivity to growth inhibition by TGF-beta. The results suggest that, in addition to structural differences, the development of functional differences in the responses of JE and OGE to TGF-beta may be associated with the formation of JE from OGE cells and the reformation of attachment after periodontal surgery.

Antibodies, Monoclonal↗
PubMed1995

Pro-transforming growth factor-alpha processing in human colon carcinoma cells: role of protein kinase C.

The human colon cancer cell lines HCT 116 (poorly differentiated) and GEO (well differentiated) express the mitogenic peptide transforming growth factor alpha (TGF-alpha). The secretion of TGF-alpha was enhanced by phorbol 12-myristate 13-acetate (PMA), indicating the possible role of protein kinase C (PKC) in the formation of mature TGF-alpha. Cells were metabolically labeled with 35S-cysteine and the formation of the mature 6 kDa TGF-alpha polypeptide from the 17 kDa pro-TGF-alpha precursor was determined. The conversion of pro-TGF-alpha was complete in 2-4 hr with the HCT 116 cells showing faster kinetics of TGF-alpha formation than GEO cells. HCT 116 cells secreted more TGF-alpha than GEO cells and the rate and extent of formation of TGF-alpha was enhanced by PMA in both cell lines. The expression of several PKC isozymes by HCT 116 and GEO cells was examined by immunoblotting. The expression of all isozymes examined was higher in HCT 116 cells compared with GEO cells. Calphostin C, an inhibitor of PKC, reduced the enzyme activity and significantly inhibited the PMA-induced secretion of TGF-alpha by both cell lines. Two agonists of PKC that act on specific PKC isozymes, thymeleatoxin and 12-deoxyphorbol 13-phenylacetate 20-acetate (dPPA), stimulated the release of TGF-alpha into the medium to the same extent as PMA. Since dPPA has been reported to stimulate PKC-beta 1 specifically, our results suggest a potential role for PKC-beta in the processing of pro-TGF-alpha by these 2 human colon carcinoma cell lines.

Colonic Neoplasms↗
PubMed1994

Involvement of protein kinase C and an elastase-like enzyme in the processing of transforming growth factor-alpha in human colon carcinoma cell lines.

Human colon carcinoma cell lines secrete transforming growth factor-alpha (TGF-alpha). Previous work indicated that the apparent m.w. of the TGF-alpha secreted by these cells ranged between 5 and 25 kDa. The more differentiated GEO cell line secreted a higher percentage of high m.w. TGF-alpha than did the poorly differentiated HCT 116 cell line. In addition, the HCT 116 cells secreted 5-fold more TGF-alpha. Treatment of HCT 116 and GEO cells with a phorbol ester (TPA) resulted in a 4-fold increase in TGF-alpha in the conditioned media of both cell types. The TPA-induced release of TGF-alpha was blocked by an inhibitor of elastase-like enzymes. This suggested a role for protein kinase C (PKC) in TGF-alpha processing in colon carcinoma cells. Direct measurement of PKC activity indicated that the HCT 116 cells (which secrete more fully processed TGF-alpha) had 10-fold more PKC activity than GEO cells. The presence of an elastase-like activity in detergent extracts and the ability of an elastase inhibitor to block the TPA-induced secretion of TGF-alpha suggests that PKC and an elastase-like enzyme are involved in the processing and secretion of TGF-alpha by human colon carcinoma cell lines.

Colonic Neoplasms↗
PubMed1993

Transforming growth factor-beta 2 is an autocrine growth inhibitory factor for the MOSER human colon carcinoma cell line.

The MOSER human colon carcinoma cell line is significantly growth inhibited by exogenous transforming growth factor-beta (TGF-beta). The secretion of TGF-beta by these cells was examined to determine if endogenous TGF-beta might also regulate MOSER cell growth. MOSER cells secreted 11 ng TGF-beta/10(6) cells, 24% of which was in the active form. Blocking antibodies specific for TGF-beta 2 stimulated growth 1.4-fold, while TGF-beta 1 specific antibodies were without effect. Treatment of MOSER cells with the differentiation agent, N,N-dimethylformamide (DMF), inhibited cell growth and resulted in an 8-fold increase in secreted TGF-beta (20% active). Only antibodies specific for TGF-beta 2 were able to reverse the growth inhibitory effect of DMF on these cells. Therefore, TGF-beta 2 acted as a negative autocrine inhibitory factor for MOSER cells and the growth inhibitory effects of DMF were mediated by the increased secretion of active TGF-beta 2.

Antibodies↗
PubMed1992

Sodium butyrate alters the response of a human colon carcinoma cell line to transforming growth factor-beta 1.

The effects of sodium butyrate (NaB) on the response of the RCA human colon carcinoma cell line to transforming growth factor-beta 1 (TGF-beta 1) were examined. NaB induced differentiation, as judged by an increase in cellular alkaline phosphatase, in the RCA cells and this differentiation was accompanied by a decreased growth rate. TGF-beta 1 did not significantly alter the growth or state of differentiation of the RCA cells. The growth rate of cells treated simultaneously with NaB and TGF-beta 1 was similar to that of control untreated cells while the alkaline phosphatase levels remained comparable to cells treated with NaB. Addition of TGF-beta 1 to cells grown in the presence of NaB resulted in a stimulation of growth. Cells pretreated with TGF-beta 1 remained sensitive to the growth inhibitory and differentiation inducing effects of NaB. These results suggest that NaB may alter the expression of proteins responsible for a stimulatory signal response to TGF-beta 1 in RCA cells.

Alkaline Phosphatase↗
PubMed1992

Response of FR3T3 cells transformed by Ha-ras oncogene and epidermal growth factor gene to differentiation induction by N,N-dimethylformamide.

Treatment of chemically transformed fibroblasts with N,N-dimethylformamide (DMF) results in the restoration of a nontransformed phenotype. In an attempt to identify more precisely the mechanisms by which DMF reverses the transformed phenotype, the effects of DMF on fibroblasts which were transformed by a single gene--specifically a synthetic epidermal growth factor (EGF) gene or the Ha-ras oncogene--were examined. The constitutive expression of either the Ha-ras oncogene or the EGF gene in FR3T3 fibroblasts resulted in cellular transformation. The effect of the differentiation-inducing agent DMF on several properties of these transformed cell lines was examined. The EGF-transfected cells were much more responsive to DMF than the ras-transfected cells. DMF treatment of the EGF-transfected cells resulted in the inhibition of anchorage-independent growth, the restoration of a normal cellular morphology and growth rate in monolayer culture, and the down-regulation of the proliferation-associated nucleolar protein B23. DMF treatment had a much slighter effect on growth of the ras-transfected cells in monolayer culture or under anchorage-independent growth conditions. The high proliferation rate of the ras-3 cells was associated with elevated expression of protein B23. The 19-3 cells, but not the ras-3 cells, expressed cell-surface fibronectin. Treatment of the ras-3 cells with DMF did not restore fibronectin expression. The binding of EGF was increased 3-fold in the EGF-transfected cells and decreased 20-fold in ras-transfected cells, but in neither case did DMF alter EGF binding. DMF treatment increased the secretion of EGF in the 2 transfected lines as well as in control cells. These results suggest that the aberrant-growth control in the EGF-transfected cells, but not in the ras-transfected cells, could be modulated by DMF and that the aberrant-growth control mechanisms were different in these 2 cell types.

Animals↗
PubMed1991

Differential role of transforming growth factor-alpha in two human colon-carcinoma cell lines.

Antibodies to epidermal-growth-factor receptor (EGF) and transforming growth factor-alpha (TGF-alpha) were used to determine the role of endogenous TGF-alpha in the growth of 2 human colon-carcinoma cell lines. Both the GEO and HCT 116 colon-carcinoma cell lines secrete similar levels of TGF-alpha and have similar numbers of low-affinity binding sites for EGF. However, the HCT 116 cells lack the high-affinity EGF binding site present on the GEO cells. The anti-EGF receptor antibodies effectively blocked the binding of 125I-EGF to the GEO and HCT 116 cell lines. Growth of the GEO cell line was inhibited 50-80% by the anti-EGF receptor and anti-TGF-alpha antibodies. When the same antibodies, in sufficient amounts to block binding of TGF-alpha to the cells, were added to the HCT 116 cell line, no effect on growth was seen. These results suggest that while the GEO cell line utilizes TGF-alpha in an autocrine manner, the TGF-alpha secreted by the HCT 116 cells apparently does not play a role in the growth of these cells.

Animals↗
PubMed1990

Characterization of the synergistic effect of insulin and transferrin and the regulation of their receptors on a human colon carcinoma cell line.

The human colon carcinoma cell line, HCT 116, can be grown in chemically defined media in the absence of exogenous growth factors. The addition of transferrin and insulin will significantly stimulate growth. The interaction of these growth factors with their receptors was studied to determine whether the synergistic action of insulin and transferrin on growth involved alterations in the growth-factor receptors. Redistribution of the transferrin receptor occurred in the presence of transferrin or transferrin plus insulin. The presence of insulin in the growth media resulted in occupation of cell-surface insulin receptors without a reduction in total insulin binding. Addition of transferrin with insulin resulted in a decrease in insulin binding to its receptor, with no alteration in receptor affinity. It appears that transferrin plays a role in regulating the insulin receptor and that this may contribute to the synergistic effect of insulin and transferrin on growth.

Carcinoma↗
PubMed1989

Up-regulation of c-myc in a transformed cell line approaching stationary phase growth in culture.

The present report describes a transformed cell line (AKR-MCA) in which the c-myc proto-oncogene is up-regulated by as much as 14-fold as cultures approach stationary phase growth. The untransformed counterpart AKR-2B cells did not exhibit such an increase in c-myc expression at high cell densities, nor did chemically transformed derivatives of another murine fibroblast cell line (C3H 10T1/2). N,N-Dimethylformamide and retinoic acid reduced c-myc levels in confluent AKR-MCA cells in association with a loss of transformed morphology, a reduction in saturation density, and the formation of a contact-inhibited monolayer at confluency. These findings suggest that the high levels of c-myc in confluent AKR-MCA cells may interfere with the normal signals involved in density-dependent growth regulation in this cell system. The effects of N,N-dimethylformamide and retinoic acid were reversible and dose-related. The half-time for the early, rapid decline in c-myc mRNA was approximately 26 min in response to N,N-dimethylformamide and 38 min in response to retinoic acid, effects which preceded the alterations in morphology and saturation densities. Activation of the latent transforming growth factor-beta in serum-free medium conditioned by confluent AKR-MCA cells, followed by its addition to preconfluent AKR-MCA cells, resulted in an up-regulation of c-myc mRNA. However, addition of serum-containing conditioned medium under similar conditions did not require prior acidification to up-regulate c-myc. Thus, active transforming growth factor-beta may be present in conditioned medium from confluent AKR-MCA cells grown in serum-containing medium, or autocrine factors other than TGF-beta may produce the confluency-associated up-regulation of c-myc and the altered density-dependent growth regulation in AKR-MCA cells.

Cell Division↗
PubMed1989

Effects of N,N-dimethylformamide and extracellular matrix on transforming growth factor-beta binding to a human colon carcinoma cell line.

Alterations in the binding of transforming growth factor-beta (TGF-beta) to the MOSER human colon carcinoma cell line caused by N,N-dimethylformamide (DMF) or extracellular matrix (ECM) were examined. DMF induced a more differentiated phenotype in the MOSER cells and resulted in a twofold increase in TGF-beta binding to the cells. This was due to an increase in receptor number with no significant alteration in the KD. The extent of increased TGF-beta binding was dependent on the dose and time of exposure to DMF. Upon removal of DMF, the receptor level returned to that of untreated cells within 6 hr. The binding of TGF-beta 1 and TGF-beta 2 to the cells was increased equally. Despite this increase in TGF-beta binding in the presence of DMF, the sensitivity of the MOSER cells to the growth inhibitory effects of TGF-beta was unaltered. Growth of the MOSER cells on ECM derived from a well-differentiated colon cell line increased the TGF-beta receptor number twofold without altering the KD. No change was observed if the MOSER cells were grown on ECM derived from a poorly differentiated cell line. While no alteration in sensitivity to TGF-beta was observed on cells grown in the presence of DMF, MOSER cells grown on the ECM derived from well-differentiated colon carcinoma cell lines were twofold more sensitive to the growth inhibitory effects of TGF-beta. These results indicated that growth conditions which resulted in a more differentiated phenotype resulted in an increase in the cellular receptors for TGF-beta.

Carcinoma↗
PubMed1989

Differential sensitivity of subclasses of human colon carcinoma cell lines to the growth inhibitory effects of transforming growth factor-beta 1.

In this study we have employed a model system comprising three groups of colon carcinoma cell lines to examine the growth-inhibitory effects of two molecular forms of transforming growth factor-beta (TGF-beta), TGF-beta 1 and TGF-beta 2. Aggressive, poorly differentiated colon carcinoma cells of group I did not respond to growth inhibitory effects of TGF-beta 1 or TGF-beta 2, while less aggressive, well-differentiated cells of group III displayed marked sensitivity to both TGF-beta 1 and TGF-beta 2 in monolayer culture as well as in soft agarose. One moderately well-differentiated cell line from group II which has intermediate growth characteristics failed to respond to TGF-beta 1 or TGF-beta 2, but the growth of two other cell lines in this group was inhibited. TGF-beta 1 and TGF-beta 2 were equally potent, 50% growth inhibition for responsive cell lines being observed at a concentration of 1 ng/ml (40 pM). Antiproliferative effects of TGF-beta 1 and TGF-beta 2 in responsive cell lines of groups II and III were associated with morphological alterations and enhanced, concentration-dependent secretion of carcinoembryonic antigen. Radiolabeled TGF-beta 1 bound to all three groups of colon carcinoma cells with high affinity (Kd between 42 and 64 pM). These data indicate for the first time a strong correlation between the degree of differentiation of colon carcinoma cell lines and sensitivity to the antiproliferative and differentiation-promoting effects of TGF-beta 1 and TGF-beta 2.

Carcinoembryonic Antigen↗
PubMed1988

Characterization of transforming growth factor-beta-resistant subclones isolated from a transforming growth factor-beta-sensitive human colon carcinoma cell line.

Previous work indicated that transforming growth factor-beta (TGF-beta) elicits proliferation-inhibitory effects in the human colon carcinoma cell line MOSER. This paper describes the isolation and characterization of spontaneously arising subclones from this TGF-beta-sensitive parental line which were relatively refractory to the inhibitory effects of TGF-beta. While the parental cell line responded to TGF-beta with an inhibition of cellular proliferation in monolayer culture and in soft agarose, an increase in extracellular fibronectin, and a down-regulation of c-myc protooncogene expression, these responses were absent or attenuated in the sublines. However, the resistant clones retained the ability to specifically bind TGF-beta. N,N-Dimethylformamide and retinoic acid, two other agents associated with induction of a partial differentiation-like response in the MOSER parental cells (similar to that elicited by TGF-beta), inhibited the monolayer proliferation of both the parental cells and the TGF-beta-resistant sublines. Thus, the refractoriness observed in the isolated clones was relatively specific for TGF-beta.

Cell Line↗
PubMed1988

Comparison of growth requirements of two human intratumoral colon carcinoma cell lines in monolayer and soft agarose.

The human colon, intratumoral subpopulations HCT 116 and HCT 116a were established in chemically defined medium supplemented with transferrin, insulin, epidermal growth factor (EGF), triiodothyronine, hydrocortisone, and sodium selenite. The responsiveness of the adapted cell lines to these growth factors was compared in anchorage-dependent and -independent assays. HCT 116 cells maintained in serum-free conditions were further adapted to growth factor deprivation, and the effects of these polypeptides were determined in anchorage-independent assays. In monolayer, HCT 116 cells adapted to grow in serum-free medium responded to transferrin but not to EGF or insulin. Similarly adapted HCT 116a cells were, however, insensitive to transferrin addition but manifested a 300 and 500% increase in growth rates with EGF and insulin, respectively. Optimal growth of HCT 116 cells was seen in the presence of insulin and transferrin, while maximum proliferation of HCT 116a cells depended on combined insulin, transferrin, and EGF. In soft agarose, both HCT 116 and HCT 116a subpopulations showed a stringent requirement for transferrin. No combination of growth factors without transferrin supported colony formation. These data suggest that (a) these colon tumor subpopulations may be subject to separate growth controls, and (b) there may be an important role for transferrin in anchorage-independent growth and possibly in the maintenance of malignant characteristics.

Cell Division↗
PubMed1988

Modulation of c-myc by transforming growth factor-beta in human colon carcinoma cells.

Previous work indicated that transforming growth factor-beta elicits proliferation-inhibitory and differentiation-like effects in the human colon carcinoma cell line MOSER. We report for the first time that the proto-oncogene c-myc is repressed in response to transforming growth factor-beta in a human colon carcinoma cell line. We also describe a subline of these cells which are relatively resistant to the transforming growth factor-beta-induced effects on proliferation in monolayer and in soft agarose, but which retain the ability to specifically bind transforming growth factor-beta. Analysis of molecular and cellular alterations in this subline may aid in elucidating the mechanism of action of transforming growth factor-beta.

Cell Division↗