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K Meflah

Publications and source records attributed to K Meflah.

At least 55 records · Page 3Linked to original sources
PubMed1997

Increase of rat colon carcinoma cells tumorigenicity by alpha(1-2) fucosyltransferase gene transfection.

Fucosylated histo-blood group antigens such as Lewis b, Lewis Y, and H accumulate in colon carcinoma and this is accompanied by a clear increase in alpha(1-2)fucosyltransferase activity, a key enzyme for the biosynthesis of these antigens. Yet the biological significance of alpha(1-2) fucosylated structures is not well defined. We have transfected a poorly tumorigenic rat colon carcinoma cell line with the human H blood group alpha(1-2)fucosyltransferase cDNA. This resulted in cell surface expression of H antigens with a concomitant decrease of sialic acid substituted and free beta-galactosides. Immunoprecipitation experiments showed that H antigens were essentially borne by variants of CD44 carrying amino acid sequences encoded by exon v6. The transfected cells showed increased motility in a wound healing assay, without changing their proliferation rates. Parental and control cells transfected with an empty vector formed small tumors that always regressed after 30 days when injected subcutaneously to syngeneic rats. In contrast, alpha(1-2)fucosyltransferase transfectants were able to form progressive tumors. Increased tumorigenicity was also visible in nude mice. These results demonstrate that alpha(1-2)fucosylated antigens contribute directly to aggressiveness of colon carcinoma cells. This could occur by altering a function of CD44 variants.

ABO Blood-Group System↗
PubMed1997

Vitamin A and apoptosis in colonic tumor cells.

The mechanism by which vitamin A prevents or delays chemical carcinogenesis remains unclear. In addition to these antimutagenic and antiproliferative activities, vitamin A seems able to induce programmed cell death. In this study, we assess the suggested role of vitamin A on the in vitro apoptosis induction in a rat colonic tumor cell line. Several concentrations of retinyl palmitate were added in the culture media. We observed cell proliferation by measuring the (3H)thymidine incorporation, cell differentiation by measuring the intestinal alkaline phosphatase expression, and apoptosis induction by DNA fragmentation and morphological evolution of adherent and floating cells. The results show that vitamin A decreases (3H)thymidine incorporation after 1 day of treatment, induces alkaline phosphatase expression, and increases the number of cells falling in apoptosis. This report confirms the role of vitamin A on the induction of cell differentiation, on the inhibition of cell proliferation and shows the vitamin A capacity to induce apoptosis. These results could be attractive to prevent development of colon cancer by vitamin A supplemented diets.

Adenocarcinoma↗
PubMed1996

Over-expression of a novel member of the immunoglobulin superfamily in Min mouse intestinal adenomas.

Germline mutations of the APC (adenomatous polyposis coli) gene lead to multiple intestinal tumors in familial adenomatous polyposis patients and in Min (multiple intestinal neoplasia) mice. Consequently, these mice provide an excellent model for familial colon cancer. We have identified an Mr approx. 66 kDa glycoprotein which is preferentially expressed at the cell surface of cell lines established from chemically induced rat colon carcinomas. Cloning of the corresponding Tage4 cDNA has revealed that this protein contains the conserved amino acids characteristic of members of the immunoglobulin gene superfamily. Here, we analyze expression of the mouse Tage4 gene in Min mouse intestinal adenomas. RT-PCR analysis allowed us to detect expression of this gene in all the mouse adenomas tested. In contrast, lower levels of Tage4 mRNA were found in the intestinal tract and barely detectable levels in other tissues of normal mice. Furthermore, Tage4 mRNA was detected in a series of mouse intestinal adenomas by in situ hybridization. A strong signal was seen in the samples analyzed.

Adenoma↗
PubMed1996

Induction of fibroblast gelatinase B expression by direct contact with cell lines derived from primary tumor but not from metastases.

During cancer progression, tumor cells interact with stromal cells. As a consequence, matrix metalloproteinases are produced that contribute to the degradation of the extracellular matrix. This study used coculture systems to investigate fibroblast interaction with three colon cancer cell lines isolated from a single patient. Cells from primary colorectal carcinoma, but not from corresponding liver or lymph node metastases, induced gelatinase B expression by fibroblasts of different tissue origin. Remarkably, direct cell-cell contact was required for this induction, which occurred at the pretranslational level (as revealed by Northern blot analysis) and was completely blocked by anti-beta1 integrin monoclonal antibody, but only partially blocked by anti-alpha5 or anti-alpha(v). Induction was also inhibited by cytochalasin D, staurosporine, or dexamethasone, suggesting the need, respectively, for an organized actin cytoskeleton, protein kinase C, and AP-1-driven gene transcription. Our data suggest that direct tumor-stromal cell contact is one inductive event involved in matrix metalloproteinase expression by stromal cells.

Animals↗
PubMed1996

Altered glycosylation of alpha(s)beta 1 integrins from rat colon carcinoma cells decreases their interaction with fibronectin.

Malignant cell transformation is generally accompanied by changes in their interactions with environing matrix proteins in a way to facilitate their migration and generate invasion. Our results show the binding of rat colon adenocarcinoma PROb cells to fibronectin strongly reduced when compared to normal rat intestine epithelial cells. This decrease was not due to the level of alpha(s)beta 1 integrins expressed at the surface of the cell line. However, beta 1- and alpha(s)-associated subunits appeared to be structurally altered as shown by immunoprecipitation followed by electrophoresis. Pulse chase experiments using 35S methionine evidenced differences in the biosynthesis of beta 1- and alpha(s)associated integrins: normal epithelial IEC18 cells required 16 h for maximal biosynthesis of the completely mature beta 1 subunit, while PROb cells did it within 4-6 h. Studies using endoglycosidases O, H, D, and N glycanase confirmed that the molecular weight alterations were due to abnormal glycosylation and suggested that alpha(s)beta 1 integrins of PROb cells could bear both mature complex and immature high mannose types while IEC18 cells borne only mature complex type oligosaccharidic chains. Treatment of both cell types with castanospermine, an inhibitor of N-glycosylation, reduced the differences observed in their adhesion to the fibronectin without significantly affecting beta 1 receptors expression at the cell surface. These results strongly suggest a role of the glycosylation of beta 1 receptors in the adhesion of rat colon adenocarcinoma PROb cells to fibronectin substrata.

Animals↗
PubMed1996

[Induction of apoptosis, in vitro and in vivo, on colonic tumor cells of the rat after sodium butyrate treatment].

Sodium butyrate (NaB) is known to induce the process of cell differentiation, particularly for epithelial colonic cells. We previously observed that treatment with NaB in association with interleukin 2 (IL2), cures 60% of peritoneal carcinomatosis induced by injection of DHDK12/TRb cells in syngenic rats [15]. In the present work, we evidenced in vitro metabolic alterations of the DHDK12/TRb cell line treated with NaB, followed by an apoptotic process. Flow cytometric analysis evidenced that the tumour cells were arrested in the G1 and G2 phases of the cell cycle for the adherent cells to the plastic. Biological analysis of cells and debris released in the culture medium were essentially apoptotic cells. Complementary, the NaB-induced apoptotic process was confirmed by the staining of the nucleus from releasing cells by Hoechst 33258 and the DNA fragmentation revealed by DNA electrophoresis. Mitochondrial activity and glucose consumption were significantly stimulated after NaB treatment, which reveal an alteration of the metabolic activity of the treated tumour cells. As a consequence, we measured a significant increase of the active TGF beta 1 production, a cytokine previously described to participate to the epithelial cell differentiation. These in vitro data were confirmed in vivo showing a significant expression of apoptotic tumour cells in NaB- or NaB/IL2-treated tumours. Thus, the present results in the rat peritoneal carcinomatosis treatment show that combination of apoptotic process induced by NaB with immunostimulation by IL2 has powerful therapeutic properties.

Animals↗
PubMed1996

[Vaccination with genetically modified IL-2 secreting cells in a rat model of colonic carcinoma].

Genetically engineered tumor cells secreting immunostimulatory molecules could facilitate the obtention of a vaccination against tumor antigens. To test this approach, we transfected genes encoding for rat and mouse IL-2 into PROb cells. These cells originate from a dimethylhydrazine induced colon carcinoma of BD IX rats. We observed an inhibition of the in vivo tumor growth directly proportional to the IL-2 secretion. An immunohistochemical analysis revealed that the tumors were infiltrated by leucocytes expressing the IL-2 receptor, suggesting their activation within the tumor. A strong delay of tumor growth was observed in rats challenged with PROb cells after a previous rejection of IL-2 secreting cells. Yet two rats out of six were completely protected. This protection is specific since rejection of PROb-IL-2 does not confer protection towards the syngeneic glioma A15A5. In addition, we could show by depletion experiments that NK/LAK, CD8, and CD4 lymphocytes were involved in the rejection of cells secreting large amounts of IL-2. Macrophages appear to be involved in the rejection process too, but also in the induction of an immune memory. Vaccination experiments using irradiated PROb IL-2 cells were performed. Only a partial protection towards a challenge with parental PROb cells could be obtained, also depending on the amount of secreted IL-2: the best protection being obtained after vaccination with cells synthesizing a small amount of IL-2. However, this protection was not superior to that obtained by coinjection of irradiated PROb cells and BCG.

Adenocarcinoma↗
PubMed1995

Glycosylation of a novel member of the immunoglobulin gene superfamily expressed in rat carcinoma cell lines.

MAb E4 recognizes a 66-kDa glycoprotein, pE4, which is a member of the immunoglobulin gene superfamily. This protein is expressed at the cell surface in rat colon and mammary carcinomas, but only in trace amounts in normal adult rat tissues. Since expression of aberrant carbohydrate structures is often associated with malignant transformation, glycosylation of pE4 was analyzed. Reactivity of lectins with pE4 suggested the absence of N-acetylneuraminic acid, terminal galactose and O-linked glycan, and the presence of N-linked glycans. Tunicamycin treatment reduced the binding of MAb E4 to cancer cells suggesting that the E4 epitope is at least partially glycosylated. Digestions with neuraminidases, O-glycosidase and peptide-N-glycosidase F confirmed these results. Pronase treatment abolished the binding of MAb E4, indicating that E4 epitope involves not only a carbohydrate determinant but also a peptide moiety. Mild periodate oxidation abolished the binding of MAb E4, indicating that non-reducing terminus carbohydrates are part of the E4 epitope. Neutral sugar analysis revealed the absence of galactose and the presence of fucose. Since fucose is sensitive to periodate oxidation, this sugar could be the carbohydrate part of the determinant recognized by MAb E4. Reactivity of lectins specific for fucose indicated the presence of alpha(1-6)-fucose on pE4.

Adenocarcinoma↗
PubMed1995

Leukaemia Inhibitory Factor derived from rat colon carcinoma cells increases host susceptibility to tumour growth.

We have tested Leukaemia Inhibitory Factor (LIF) production by 12 rat colon tumour clones isolated from a single cell line that display various degrees of tumorigenicity. A highly significantly relationship was found between levels of soluble LIF produced by the clones and their in vivo tumorigenicity. Such results suggested a role for LIF as a tumour facilitating agent. To test this hypothesis, the highly tumorigenic and LIF producing PROb clone was transfected with the LIF cDNA in antisense orientation in order to decrease LIF production. Conversely, REGb, a low LIF producer that is rejected by syngeneic animals, as well as nude mice, was transfected with the LIF cDNA to increase its production. PROb cells transfected with antisense cDNA were shown to have decreased LIF production along with decreased tumorigenicity. LIF-transfected REGb cells expressing high LIF levels still regressed in syngeneic rats, but could form progressive tumours in nude mice. We did not detect LIF receptors on PROb or REGb cells and their in vitro proliferation was not modified by the addition of exogenous LIF. Therefore, LIF was not an autocrine growth regulator for PROb and REGb cells. Instead, LIF appears to facilitate in vivo tumour growth, without being an immunosuppressive factor sufficient on its own to allow growth of immunogenic cells in fully immunocompetent hosts.

Adenocarcinoma↗
PubMed1995

Immunization against a rat colon carcinoma by sodium butyrate-treated cells but not by interleukin 2-secreting cells.

BACKGROUND & AIMS: Vaccination of patients with colon cancer with irradiated autologous tumor cells and bacille Calmette-Guérin (BCG) was reported to augment mean survival. It was recently observed that a local treatment combining recombinant interleukin 2 and the differentiation agent sodium butyrate cured rats with colon cancer peritoneal carcinomatosis. To optimize vaccination protocols, the comparison of the efficacy of irradiated tumor cells mixed with BCG with that of interleukin 2-gene-transfected cells and of tumor cells pretreated with sodium butyrate was performed. METHODS: The poorly immunogenic rat colon carcinoma cells PROb were used in a vaccination assay. Interleukin 2-transfected PROb cells, either proliferating or irradiated, were used. The efficiency of irradiated PROb cells mixed with BCG, of interleukin 2-transfected cells, or of cells pretreated with sodium butyrate was tested. RESULTS: Vaccination with irradiated parental cells and BCG did not provide protection. Irradiated interleukin 2-transfected cells were poorly efficient in the vaccination assay. Conversely, vaccination with irradiated parental cells pretreated with sodium butyrate before injection provided good protection. CONCLUSIONS: Interleukin 2-secreting cells efficiently vaccinated animals when injected while replicating but not after irradiation. Conversely, sodium butyrate pretreatment provided a simple and efficient vaccination scheme that generated a long-term immune memory and allowed the use of irradiated cells.

Animals↗
PubMed1995

The expression of carbohydrate blood group antigens correlates with heat resistance.

Recent data indicate that cells may resist heat shock via more than one route: heat shock protein synthesis and other still ill-defined mechanisms. We investigated this phenomenon using four types of cells derived from a single rat colon carcinoma: clones REGb and PROb; PRO A+, a glycosylation variant of PROb selected for its high expression of blood group A antigen; and Ph8, a thermoresistant variant of PROb selected by repeated sublethal heat treatments. Basal heat resistance was clearly associated with the level of cell surface expression of blood group H and A antigens. Biosynthesis of these carbohydrate structures requires two glycosyltransferases, H and A enzymes, whose activities are also correlated with basal heat resistance. In addition, heat sensitive REGb cells were rendered more resistant by transfection with the gene encoding for H enzyme, allowing expression of H antigen. Thus, these terminal glycosylations could play a role as cellular protectors against heat treatment. Blood group carbohydrate antigens were mainly located on O-linked carbohydrate chains of a major glycoprotein of 200 kDa and to a lesser extent on N-linked chains. Only trace amounts were present as glycolipids.

ABO Blood-Group System↗
PubMed1994

The role of transforming growth factor beta 1 in the fibroblastic reaction associated with rat colorectal tumor development.

Many tumors are surrounded by a highly fibrous stroma composed of fibroblasts and extracellular matrix. This desmoplastic response has been suggested to both inhibit and favor tumor progression. The present study deals with the effects of tumor cells on the fibroblastic reactions they cause and relates this to progression or regression of tumors. Two rat colon carcinoma cell lines, one which develops progressive tumors when injected s.c. in syngeneic animals (PROb cell line) and the other which develops regressive tumors in similar conditions (REGb cell line), were compared by the fibroblastic reaction which they cause. Comparative histological analysis of progressive and regressive tumors developed by the two cell lines showed a significant but opposite response of fibroblastic compartment. The progressive tumor nodules were observed to grow within a loose tissue, whereas the regressive tumor cells were surrounded by a fibrous capsule. Immunohistological labelings revealed the presence of alpha-smooth muscle actin-positive myofibroblasts during the tumor expansion, while these specific cells disappeared during the tumor regression. Immunostainings of transforming growth factor beta 1 showed an increasing staining of the progressive tumor cells during tumor development but a slight expression by tumor cells and stroma during the tumor regression. This growth factor was demonstrated to facilitate initial steps of the tumor progression by addition of active transforming growth factor beta 1 at the time of s.c. injection of PROb cells in syngeneic rat models. In vitro experimental analysis with the use of neutralizing antibody showed that active transforming growth factor beta produced by the progressive cells inhibited fibroblast proliferation and facilitated their differentiation into myofibroblasts. Since the number of myofibroblasts increased with time in progressive tumors, their presence may constitute a potential growth advantage for tumor growth. In contrast, our results indicated involvement of platelet-derived growth factor-like protein(s) in fibroblast proliferation under the control of regressive cells and the presence of an important sheath of alpha-smooth muscle actin-negative fibroblasts in regressive tumors may support a role for this growth factor in vivo. Thus, the ability of tumor cells to produce or induce the production of transforming growth factor beta or platelet-derived growth factor may give rise to a specific fibroblast reaction, which in turn may determine consequent tumor evolution.

Actins↗
PubMed1994

H blood group antigen carried by CD44V modulates tumorigenicity of rat colon carcinoma cells.

Expression of carbohydrate ABH blood group antigens is oncodevelopmentally regulated and their presence on tumor cells constitutes a prognostic factor. However, it is not clear whether they directly affect tumor behavior. Using a rat model of colon carcinoma, we previously observed an association between the presence of H blood group antigens and tumorigenicity in syngeneic animals. In the present study, we show by immunoprecipitation experiments that cell surface H blood group antigens of a highly tumorigenic clone (PROb) are essentially carried by splice variants of the CD44 molecule containing exon V6. PROb cells were then transfected with an antisense fragment of the gene coding for a rat alpha (1-2)fucosyltransferase. This enzyme allows synthesis of H antigens from various beta-galactoside precursors. Transfected subclones of PROb cells were obtained which had significantly decreased enzymatic activity and H antigenic cell surface levels. In contrast, no such changes were observed in control cells transfected with either the empty vector or with a sense fragment of the gene. Compared to controls, the antisense-transfected cells were far less tumorigenic in syngeneic animals. These results show that H blood group antigens at the surface of PROb colon carcinoma cells contribute to tumor progression. The presence of the fucosylated structures on CD44 could modulate the functions of this adhesion molecule.

ABO Blood-Group System↗
PubMed1994

Expression of the 100-kda glucose-regulated protein (GRP100/endoplasmin) is associated with tumorigenicity in a model of rat colon adenocarcinoma.

Resistance to glucose starvation and expression of glucose-regulated proteins (grp) were studied in a model of rat colon carcinoma. In this model, various clones originating from the same parental tumor showed distinct tumorigenic potential in syngeneic hosts. Some clones were tumorigenic while others were rejected by an immune-based mechanism. It appeared that the more tumorigenic clones were more resistant to glucose starvation and able to synthesize larger amounts of grp100 than non-tumorigenic clones in response to either glucose starvation or tunicamycin treatment. In contrast, there was no difference in the induced levels of synthesis of grp78 between tumorigenic and non-tumorigenic clones. These results suggest that the ability of cells to synthesize large amounts of the stress protein grp100 might allow them to resist marginal conditions imposed by fully immunocompetent hosts, thus conferring greater tumorigenicity.

Adenocarcinoma↗
PubMed1994

Chemotherapeutic inhibition of erb-B2 oncogene expression on a non-small-cell cancer line (NSCLC-N6) by marine substances.

The inhibitory effect of natural substances of marine origin on the erb-B2 oncogene of a human NSCLC-N6 line was demonstrated in vitro by simultaneous study of the expression of the gene and its product, using respectively an erb-B2 specific probe and an anti-c-erb-B2 polyclonal antibody. Preliminary results indicate inhibition ranging from 17-77% of oncogene expression and from 77-90% of product expression. The fact that substances of this type, with different chemical structures, have the common ability to induce terminal differentiation in an experimental model after irreversible blockade in G1 phase suggests a relationship between the inhibition of certain oncogenes and terminal differentiation.

Acetamides↗
PubMed1993

Over-expression of the S13 ribosomal protein in actively growing cells.

In order to define biological markers of aggressiveness, 2 rat colon-carcinoma cell lines differing by their tumorigenicity were used to clone genes over-expressed in colon carcinoma as compared with normal epithelial cells. A progressive rat colon-carcinoma clone (PROb) cDNA library was hybridized with 32P-cDNA synthesized from mRNA prepared from these PROb cells, or from regressive cells (REGb) derived from the same tumor. Several clones were isolated after the initial screening. The specificity of each clone was confirmed by RNA blotting. One of these (B9) was found to hybridize to an mRNA 30-fold more abundant in PROb cells than in normal adult rat colon, and was therefore selected for further study. No gene amplification was detected by Southern blot analysis, indicating that the difference in mRNA content was most likely due to an increased transcription of this gene. Sequencing of the cDNA revealed approximately 98% homology with the rat S13 ribosomal protein. The expression level of this gene was determined in a series of rat cell lines with different growth rates. A good correlation was found between these 2 parameters. Our data suggest that the S13 ribosomal-protein gene can be used to evaluate the growth rate of tumor cells, which might be correlated with their aggressiveness. In an initial trial experiment, S13 ribosomal-protein mRNA was detected in a series of human colorectal tumors by in situ hybridization. A strong signal was seen in the 4 tumors analyzed.

Adenocarcinoma↗
PubMed1993

Analysis of the relationship between stage of differentiation and NK/LAK susceptibility of colon carcinoma cells.

NK and LAK cells which are able to lyse tumor target cells in an MHC-unrestricted manner are not equally effective against targets of the same nature. In the case of colorectal tumors, some cells are highly sensitive, whereas others are resistant to NK and can even be quite resistant to LAK-mediated lysis. In the present paper, we tried to correlate the stage of differentiation of 17 human colorectal tumor cell lines with their NK- or LAK-cell susceptibility. It was observed that NK cells killed colorectal target cells independently from their stage of differentiation defined according to histopathological criteria from xenografting in nude mice. NK susceptibility was not correlated either with in vitro-defined criteria of differentiation, such as cell polarity and morphology, brush-border enzyme expression and CEA production. A LAK-resistant HT-29 sub-line (HT-29 LAK) was selected which could not be distinguished from HT-29 in terms of features of differentiation. It was further observed that HT-29 Glc-/+ cell line, a highly differentiated enterocytic-like variant of HT-29, obtained after glucose starvation, was killed by LAK cells as efficiently as the moderately differentiated parental HT-29, and that Caco-2 cells, which differentiate spontaneously after confluence in standard culture conditions, were equally sensitive to NK-mediated lysis whatever their stage of differentiation. In contrast, HT29 MTX10(-5), a highly differentiated mucus-secreting variant of HT29 obtained by methotrexate selection, was much more resistant to LAK cells than parental HT29 cells.

Animals↗