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At least 595 records · Page 33Linked to original sources
PubMed1999

Osmotic shrinkage activates nonselective cation (NSC) channels in various cell types.

Osmotic cell shrinkage activates a nonselective cation (NSC) channel in M-1 mouse cortical collecting duct cells (Volk, Frömter & Korbmacher, 1995, Proc. Natl. Acad. Sci. USA 92: 8478-8482). To see whether shrinkage-activated NSC channels are an ubiquitous phenomenon, we tested the effect of hypertonic extracellular solution on whole-cell currents of HT29 human colon carcinoma cells, BSC-1 renal epithelial cells, A10 vascular smooth muscle cells, and Neuro-2a neuroblastoma cells. Addition of 100 mm sucrose to an isotonic NaCl bath solution induced cell shrinkage of HT29 cells as evidenced by a decrease in cell diameter from 18 +/- 1 microm to 12 +/- 1 microm (n = 13). Upon cell shrinkage whole-cell currents of HT29 cells increased within 8 +/- 1 min by about 30-fold (n = 13). Cell shrinkage and current activation were reversible upon return to isotonic solution. Replacement of bath Na+ by K+ or Li+ had almost no effect on the stimulated inward current. In contrast, replacement by N-methyl-d-glucamine (NMDG) completely abolished it and shifted the reversal potential from -4.5 +/- 0.7 mV to -57 +/- 4.1 mV (n = 10). Thus, the stimulated conductance is nonselective for alkali cations but highly selective for cations over anions with a cation-to-anion permeability ratio of about 13. Flufenamic acid (100 microm) inhibited the stimulated current by 84 +/- 4.7% (n = 8). During the early phase of hypertonic stimulation single-channel transitions could be detected in whole-cell current recordings, and a gradual activation of 12 and more individual channels with a single-channel conductance of 17.6 +/- 0.9 pS (n = 4) could be resolved. In analogous experiments similar shrinkage-activated NSC channels were also observed in BSC-1 renal epithelial cells, A10 vascular smooth muscle cells, and Neuro-2a neuroblastoma cells. These findings indicate that shrinkage-activated NSC channels are an ubiquitous phenomenon and may play a role in volume regulation.

Animals↗
PubMed1999

Expression and regulation by interferon-gamma of the membrane-bound complement regulators CD46 (MCP), CD55 (DAF) and CD59 in gastrointestinal tumours.

The membrane-bound complement inhibitors CD46 (membrane cofactor protein), CD55 (decay-accelerating factor) and CD59 (protectin) protect tumour cells against lysis by activated complement. In this study, a total of 14 (3 gastric, 3 colonic and 8 pancreatic) gastrointestinal tumour cell lines were examined for the expression of CD46, CD55 and CD59 with respect to the regulatory efficacy of interferon-gamma (IFN-gamma). The effects of IFN-gamma on mRNA and protein expression levels of CD46, CD55 and CD59 were evaluated by Northern blot hybridisation, RT-PCR, flow cytometry and immunostaining. In unstimulated cell lines, CD46 and CD59 transcripts were expressed at comparable levels, whereas the basal expression of CD55 mRNA was heterogeneous. The complement inhibitor proteins were detected in all cell lines using specific antibodies. Additional immunohistochemical stainings of gastrointestinal tissue specimens supported these findings. IFN-gamma evoked a weak induction of certain transcripts in a subset of the cell lines. Upregulation of protein expression was only observed in HT29 cells for CD55 and CD59 and was accompanied by a marked increase of the corresponding transcripts. We conclude that membrane-bound complement inhibitors are broadly expressed in gastrointestinal tumour cells and vary in their susceptibility to IFN-gamma. Thus, they may be involved in tumour escape mechanisms in gastric, pancreatic and colorectal cancer.

Antigens, CD↗
PubMed2002

Serotypes, virulence factors, antibiotic sensitivity, beta-lactamase activity and plasmid analysis of Salmonella from children with diarrhea in Tripoli (Libya).

A total of 21 Salmonella strains isolated in Libya (16 from children with diarrhea and 5 from healthy controls) were serotyped and studied for their cell invasive ability, production of cytotoxin, antibiotic susceptibility, beta-lactamase activity and plasmid profiles. Eight different serotypes of Salmonella were identified: 6 S. saintpaul, 4 S. wien (1 from control), 2 S. newport, 2 S. muenchen (1 from control), 2 S. typhimurium (1 from control), 2 S. hadar (1 from control), 2 S. reading (1 from control), 1 S. kottbus. Twenty (95%) were positive in the invasiveness assay using HeLa cells, and all (100%) were negative for cytotoxin production in HT29 cells. More than 40% were resistant to ampicillin, cefalexin, cefamandole, cefoperazone, chloramphenicol, gentamicin, mezlocillin and trimethoprimsulphamethoxazole and 100% were susceptible to the new quinolones. Most (67%) of the strains harbored plasmids and 43% produced beta-lactamase. A strong association was observed between the presence of more than one plasmid, beta-lactamase activity, and multiple-resistance to antimicrobial agents and serotypes S. saintpaul and S. wien. Curing experiments with acridine orange showed that 2 plasmids (33 and 1.4 megadaltons) might be responsible for the resistance to chloramphenicol and gentamicin. The present study demonstrated that multiple-resistant salmonellae are widespread in Libya and the resistance is mainly plasmid mediated.

Anti-Bacterial Agents↗
PubMed1983

Subcellular distribution, synthesis, and release of carcinoembryonic antigen in cultured human colon adenocarcinoma cell lines.

The content of carcinoembryonic antigen (CEA) and its subcellular distribution were studied in nine human colon carcinoma cell lines. A great variation in CEA content was found among different colon cancer cell lines. Well-differentiated colon cancer cell lines (LS174T and SKCO-1) contained the highest CEA activity which was 35 to 60 times greater than that of less well-differentiated cells (SW620, SW480, and HRT18). More than 80% of the CEA was associated with the cell membrane in all nine cell lines. With increasing cell density, the CEA content per cell was found to increase in SW1116, HCT8, and HCT48 cells, while no change was observed in SW620, HRT18, and HT29 cells. In SW480, LS174T, and SKCO-1 cells, CEA content actually decreased with increasing cell density. Investigation of the synthesis of CEA in cells and its release into the medium over an 8-day period showed that the rate of CEA synthesis at maximum cell density for LS174T and SKCO-1 cells decreased to 15 and 50% of that at low cell density, respectively. In contrast, the rate of CEA release into medium by these two cell lines was higher at maximum than at low cell density. For HCT48 cells, the increased rate of CEA synthesis with increasing cell density markedly elevated cellular CEA levels. These observations were confirmed by studying the rate of incorporation of N-acetyl[3H]-glucosamine into cellular CEA in LS174T and HCT48 cell lines. The rate of incorporation of radioactivity into CEA was greater during the exponential phase of growth than during the stationary phase for LS174T, while the opposite was observed with HCT48 cells. This study indicated that there is a great variation in CEA content among different human colon cancer cell lines and that it is associated predominantly with the membrane fraction. The rate of synthesis and release of CEA also varied among different cell lines. The growth phase had a varied effect on the CEA content, the rate of synthesis, and the release of CEA in these human colon cancer cell lines.

Adenocarcinoma↗
PubMed1984

A monoclonal antibody against a CEA-related antigen expressed on HT29 colon tumour cells.

A new monoclonal antibody, that binds to CEA and with low cross-reactivity with NCA, has been raised to an antigen expressed on HT29 colon tumour cells. This antibody (H58) reacts strongly with high-molecular-weight protein (50 X 10(4)) isolated from a crude plasma membrane preparation of HT29 cells as well as binding to purified CEA of molecular size (20 X 10(4)) isolated both from those cells and liver metastases of colon tumour. H58 binds to an epitope sterically unrelated to the binding site of the previously described anti-CEA monoclonal antibody MA/1 and our routine anti-CEA polyclonal serum PKIG. Augmented binding of antibody to either the cell membrane preparation or conventional CEA can be achieved using a mixture comprising equal weights of specific immunoglobulin from H58 and MA/1. The value of solid-phase binding assays using microtitre plates for selecting potentially useful antibodies is discussed.

Animals↗
PubMed1996

Expression of the integrin alpha 5 subunit in HT29 colon carcinoma cells suppresses apoptosis triggered by serum deprivation.

It is clear that certain integrins can regulate the growth of tumors, probably by contributing to signal transduction processes. In the present study we have used HT29 human colon carcinoma cells stably transfected with human cDNA for the integrin alpha 5 subunit and studied the effects of alpha 5 expression on the induction of apoptosis. We observe that apoptosis can be triggered in HT29 cells by removal of serum and that this process can be suppressed by the stable expression of full-length integrin alpha 5 subunits. While the mechanism underlying this effect is still unclear, these observations suggest that the alpha 5 beta 1 integrin plays an important role in modulating tumor cell responses to growth factors and nutrients.

Antigens, CD↗
PubMed1998

Convergent effects of growth factors, hormones, and fibronectin are necessary for the enterocyte differentiation of a colon adenocarcinoma cell line (HT29-D4).

The aim of this work was to show in serum-free medium a convergent effect of physiological factors and extracellular matrix proteins on the differentiation process of enterocytes by taking as a model the HT29-D4 clone that has the feature of differentiating when subcultured in fetal bovine serum glucose-free medium. We show that triiodothyronine (T3) as well as insulin promotes limited cell growth and differentiation, whereas fibronectin or bovine serum albumin (BSA) induces cell growth and a low level of differentiation. However, insulin, T3, fibronectin, and BSA together with epidermal growth factor and transferrin promoted satisfactory growth and enterocyte morphology with epithelial electrophysiological properties in HT29-D4 cells. With these factors adequate protein targeting was achieved since cells apically expressed the carcinoembryonic antigen, and basolaterally transferrin and insulin receptors, beta 1 and alpha v beta 6 integrins, talin, vinculin, and focal adhesion kinase (FAK). Talin, vinculin, FAK, and alpha v beta 6 integrin, the fibronectin receptor, were clustered in focal contacts, which agrees with a possible role of fibronectin in final cell growth, the latter process mediating the final phase of differentiation. This level of differentiation can be maintained for a long time. Thus HT29-D4 cells appear to be a suitable model to study the implication of integrins in the differentiation process of human enterocytes.

Adenocarcinoma↗
PubMed1994

Modifications of the anti-oxidant metabolism during proliferation and differentiation of colon tumor cell lines.

The anti-oxidant metabolism was studied at different times after sub-culture in 2 colon cell lines previously characterized for their growth and differentiation properties. The HT29 cell line is mainly composed of proliferative and undifferentiative cells, while the derived 5-fluorouracil (FUra)-adapted cells undergo growth-dependent differentiation, which is complete at post-confluence. In the 2 cell lines, all the anti-oxidant parameters studied appeared to be related to proliferation, with increased activity of superoxide dismutase (SOD) 1 and 2, catalase (CAT), glutathione peroxidase (GPX), glutathione reductase (GSR), and glutathione transferase (GST), and decreased glucose-6-phosphate dehydrogenase (G6PD) activity and glutathione content, in parallel with slowing down of proliferation. At post-confluence, these metabolic parameters remained stable, except for GPX activity, which continued to increase, and CAT activity, which decreased. The amounts of SOD1, SOD2 and CAT immunoreactive proteins, estimated by Western blotting, appeared to be correlated to their respective enzymatic activities. SOD1, CAT and GST activity and glutathione content, which remained at similar levels in the 2 cell lines for all times studied, appeared unrelated to the differentiation process. GSR and GPX activity, which was lower in FUra-adapted than in parental cells only at post-confluence, could be considered as markers of differentiated cells. The higher SOD2 and lower G6PD activity observed in FUra-resistant cell in comparison with parental cells at all times after sub-culture could be characteristic both of differentiative and of differentiated cells. Interestingly, cytogenetics have previously indicated that deletions of the long arm of chromosome 6, which carry the gene for SOD2, were frequently observed in parental but not in FUra-adapted cells. These results demonstrate that modifications of the anti-oxidant metabolism occur in relation with proliferation and differentiation, and suggest a particular role for SOD2 in these cellular processes.

Antioxidants↗
PubMed2004

2-amino-O4-benzylpteridine derivatives: potent inactivators of O6-alkylguanine-DNA alkyltransferase.

2-amino-O4-benzylpteridine (1), 2-amino-O4-benzyl-6,7-dimethylpteridine (2), 2-amino-O4-benzyl-6-hydroxymethylpteridine (4), 2-amino-O4-benzylpteridine-6-carboxylic acid (5), 2-amino-O4-benzyl-6-formylpteridine (6), and O4-benzylfolic acid (7) are shown to be as potent or more potent inactivators of the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (alkyltransferase) in vitro than O6-benzylguanine, the prototype alkyltransferase inactivator currently in clinical trials. Additionally, the negatively charged (at physiological pH) inactivators 2-amino-O4-benzylpteridine-6-carboxylic acid (5) and O4-benzylfolate (7) are far more water soluble than O6-benzylguanine. The activity of O4-benzylfolic acid (7) is particularly noteworthy because it is roughly 30 times more active than O6-benzylguanine against the wild-type alkyltransferase and is even capable of inactivating the P140K mutant alkyltransferase that is resistant to inactivation by O6-benzylguanine. All the pteridine derivatives except 2-amino-O4-benzylpteridine-6-carboxylic acid are effective in enhancing cell killing by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). However, the effectiveness of O4-benzylfolate as an adjuvant for cell killing by BCNU appears to be a function of a cell's alpha-folate receptor expression. Thus, O4-benzylfolate is least effective as an adjuvant in A549 cells (which express little if any receptor), is moderately effective in HT29 cells (which express low levels of the receptor), but is very effective in KB cells (which are known to express high levels of the alpha-folate receptor). Therefore, O4-benzylfolic acid shows promise as an agent for possible tumor-selective alkyltransferase inactivation, which suggests it may prove to be superior to O6-benzylguanine as a chemotherapy adjuvant.

Cell Line, Tumor↗
PubMed2004

Antiproliferative effects of ZD0473 (AMD473) in combination with 5-fluorouracil or SN38 in human colorectal cancer cell lines.

PURPOSE: ZD0473 (AMD473) [cis-amminedichloro(2-methylpyridine) platinum(II)] is a novel platinum agent of proven activity in vitro against a variety of human tumor-derived cell lines even with intrinsic or acquired resistance to CDDP. The aim of this study is to provide the basis for a rational design of ZD0473-based combination in colon cancer. EXPERIMENTAL DESIGN: We evaluated the cytotoxic effect of ZD0473 administered alone or in combination with 5-Fluorouracil (5FU) or SN38 in a panel of sensitive and 5FU-resistant colorectal cell lines (HT29/HT29-5FUR and LoVo/LoVo-5FUR). We analyzed four sequential schedules of administration: ZD0473 --> 5FU, 5FU --> ZD0473, ZD0473 --> SN38 and SN38 --> ZD0473. MTT-assay and isobologram analyses were performed to determine the synergism/antagonism. RESULTS: The pattern of response towards ZD0473, administered as single agent, was similar in all cases and independent of the 5FU-resistance phenotype (IC50 from 48.1 to 76.6 microM) and/or p53 status. No differences in sensitivity to ZD0473 alone or in combination were observed between DNA-mismatch repair-proficient (HT29/HT29-5FUR) and -deficient (LoVo/LoVo-5FUR) cells. ZD0473 administered prior to 5FU leads to synergistic/additive effect in all cell lines, while the 5FU --> ZD047 schedule was only synergistic in HT29 cells. Exposure to ZD0473 prior to SN38 leads to a synergistic/additive schedule in LoVo/LoVo-5FUR cells, while SN38 --> ZD0473 schedule was only synergistic in parental cell lines. CONCLUSIONS: The combinations of ZD0473 and 5FU or SN38 have shown to be active in sensitive and 5FU-resistant colorectal cell lines when a correct schedule of administration is applied. These results may be further exploited to promote new schedules of administration for advanced colorectal cancer treatment.

Antineoplastic Agents↗
PubMed2000

Novel polymerase chain reaction approach for full-coding p53 mutation detection in microdissected archival tumors.

Evidence suggests that up to 25% of p53 mutations are outside of exons 5-8 and that insertions, deletions, and polymorphic sites in the p53 gene may play a significant role in the process of carcinogenesis. A novel polymerase chain reaction (PCR) approach for the analysis of the entire p53 coding and splice site regions from microdissected, formalin-fixed, paraffin-embedded tumor tissues has been developed which allows multiple genetic analyses to be performed from one primary amplification reaction. The method was initially evaluated using well-characterized cell lines. In addition to confirming the published p53 mutations for HT29, Molt 4, A431, and HN5, a 16 base pair (bp) duplication within intron 3 was detected in both the A431 and HT29 cell lines. Analysis of archival samples of ovarian cancer identified the same 16-bp duplication and coding region variations. In all samples, using GenBank submission U94788 as a reference, a C-insertion was detected at nucleotide positions 11818 and 11874 within intron 2. At nucleotide position 14168, within intron 7, a T-to-G base change was found. This novel PCR approach has the potential to reduce the amount of clinical material required by up to 95%, thus facilitating retrospective studies on archival tumor collections. Furthermore, a wider analysis of the p53 gene, including splice sites and intronic regions, may yield additional information regarding cancer predisposition, response to therapy, and progression.

Base Sequence↗
PubMed2003

Purification, localization, and expression of human intestinal alkaline sphingomyelinase.

Sphingomyelin (SM) metabolism in the gut may have an impact on colon cancer development. In this study, we purified alkaline sphingomyelinase (alk-SMase) from human intestinal content, and studied its location in the mucosa, expression in colon cancer, and function on colon cancer cells. The enzyme was purified by a series of chromatographies. The molecular mass of the enzyme is 60 kDa, optimal pH is 8.5, and isoelectric point is 6.6. Under optimal conditions, 1 mg of the enzyme hydrolyzed 11 mM SM per hour. The properties of the enzyme are similar to those of rat intestinal alk-SMase but not to those of bacterial neutral SMase. Immunogold electronmicroscopy identified the enzyme on the microvillar membrane in endosome-like structures and in the Golgi complexes of human enterocytes. The expression and the activity of the enzyme were decreased in parallel in human colon cancer tissues compared with the adjacent normal tissue. The enzyme inhibited DNA biosynthesis and cell proliferation dose dependently and caused a reduction of SM in HT29 cells. Intestinal alk-SMase is localized in the enterocytes, down-regulated in human colon cancer, and may have antiproliferative effects on colon cancer cells.

Apoptosis↗
PubMedDate not supplied

P27KiP1 overexpression inhibits the growth and doxorubicin sensitivity of HT29 human colon cancer cells in vivo.

We have previously shown that p27KiP1 plays a role in the tumor cell resistance of HT29 confluent monolayers to cytotoxic drugs in vitro. To determine whether p27KiP1 was a resistance factor to cytotoxic drugs in vivo we tested the effect of doxorubicin on p27KiP1-overexpressing HT29 tumors in nude mice. In this study we show that ectopic overexpression of p27KiP1 in HT29 human colon cancer cells decreases their tumorigenicity in vivo in nude mice. This decreased tumor growth was associated with increased p27KiP1 protein expression, studied by Western blotting in tumor extracts. Interestingly, the overexpressing-p27KiP1 tumors were significantly more resistant to intraveneous doxorubicin treatment than the control tumors. These results indicate that p27KiP1, which delays tumor growth could also increase tumor resistance to cytotoxic drugs in vivo.

Animals↗
PubMed2005

Cell-surface targeting of alpha2-adrenergic receptors -- inhibition by a transport deficient mutant through dimerization.

We previously demonstrated that the alpha2B-adrenergic receptor mutant, in which the F(x)6IL motif in the membrane-proximal carboxyl terminus were mutated to alanines (alpha2B-ARm), is deficient in export from the endoplasmic reticulum (ER). In this report, we determined if alpha2B-ARm could modulate transport from the ER to the cell surface and signaling of its wild-type counterpart. Transient expression of alpha2B-ARm in HEK293T cells markedly inhibited cell-surface expression of wild-type alpha2B-AR, as measured by radioligand binding. Subcellular localization demonstrated that alpha2B-ARm trapped alpha2B-AR in the ER. The alpha2B-AR was shown to form homodimers and heterodimers with alpha2B-ARm as measured by co-immunoprecipitation of the receptors tagged with green fluorescent protein and hemagglutinin epitopes. In addition to alpha2B-AR, the transport of alpha2A-AR and alpha2C-AR to the cell surface was also inhibited by alpha2B-ARm. Furthermore, transient expression of alpha2B-ARm significantly reduced cell-surface expression of endogenous alpha2-AR in NG108-15 and HT29 cells. Consistent with its effect on alpha2-AR cell-surface expression, alpha2B-ARm attenuated alpha2A-AR- and alpha2B-AR-mediated ERK1/2 activation. These data demonstrated that the ER-retained mutant alpha2B-ARm conferred a dominant negative effect on the cell-surface expression of wild-type alpha2-AR, which is likely mediated through heterodimerization. These data indicate a crucial role of ER export in the regulation of cell-surface targeting and signaling of G protein-coupled receptors.

Alanine↗
PubMed1998

Upregulation of CASP genes in human tumor cells undergoing etoposide-induced apoptosis.

Caspases are aspartate-specific cysteine proteases that play a pivotal role in drug-induced cell death. We designed RT-PCR assays to analyse the expression of CASP-3, CASP-4, CASP-6 and the long and short isoforms of CASP-2 genes in human cells. These genes heterogeneously coexpress in leukemic cell lines and bone marrow samples from patients with de novo acute myelogenous leukemia at diagnosis. Treatment of U937 and HL60 leukemic cells and HT29 colon carcinoma cells with the topoisomerase II inhibitor etoposide upregulates CASP-2 and CASP-3 genes in these cells before inducing their apoptosis. This effect of etoposide is not observed in K562 cells and bcl-2-transfected U937 cells which are less sensitive to drug-induced apoptosis. Nuclear run-on experiments demonstrate that etoposide increases CASP gene transcription in U937 cells, an effect that is prevented by Bcl-2 overexpression. Upregulation of CASP genes is associated with an enhanced synthesis of related procaspases that precedes the appearance of apoptosis markers including caspase-3 activation, poly(ADP-ribose) polymerase cleavage and internucleosomal DNA fragmentation. These results suggest that the ability of tumor cells to upregulate CASP-2 and CASP-3 genes in response to cytotoxic drugs could be predictive of their sensitivity to drug-induced apoptosis.

Apoptosis↗
PubMed2006

Increased radiosensitivity in cells of two human cell lines treated with bystander medium from irradiated repair-deficient cells.

Radiation-induced bystander factors have been shown to be more toxic if they are from medium harvested from irradiated repair-deficient cells. The aim of this study was to test the hypothesis that the radiosensitivity of repair-proficient cells can be increased by exposing them to medium-borne factors harvested from sensitive cells and vice versa. Cells from a mismatch repair (MMR)-deficient cell line (Raji 10) with a sensitive response to radiation or the wild-type parent cell line were irradiated to 0.5 Gy gamma rays and then monitored for growth rate in their own medium or in the alternative conditioned medium. In other experiments, cells or conditioned medium were added to reporter cells (HPV-G, which are relatively sensitive keratinocytes, or highly radioresistant HT29 cells). The subsequent responses of the two cell lines to a 0.5-Gy dose of (60)Co gamma rays were measured. The results show that prior exposure of resistant cells to medium from irradiated sensitive cells reduced the clonogenic survival of the subsequently irradiated resistant cells. The reverse is also true. Measurement of the apoptosis index and BCL2 expression confirmed that the harvested medium was capable of modulating apoptosis after irradiation. This may have important applications in tumor therapy and also in the understanding of mechanisms involved in induction of adaptive responses.

Apoptosis↗
PubMed2003

Vasoactive intestinal peptide stimulates proliferation in HT29 human colonic adenocarcinoma cells: concomitant activation of Ras/Rap1-B-Raf-ERK signalling pathway.

The vasoactive intestinal peptide (VIP) has been shown to regulate cell proliferation and differentiation in many cell types. We previously reported that this neuropeptide inhibited proliferation in HT29 adenocarcinoma cells cultured in serum-containing medium. In addition, it has been demonstrated that VIP induced a potent stimulation of intracellular cAMP production in these cells cultured either in the absence or in the presence of serum. We also demonstrated that VIP induced phosphorylation of the small GTPase Rap1 in these cancerogenous cells. In the present study, the effects of VIP on the proliferation of HT29 cells cultured in the absence of growth factors and various concomitant signalling events were investigated. Under serum-free conditions VIP stimulates HT29 cell proliferation and induced a time- and concentration-dependent ERK activation. Furthermore, VIP induced the activation of the small GTPase Rap1 and of a 95 kDa isoform of the serine/threonine kinase B-Raf. Ras GTPase is also activated in VIP-stimulated cells. We hypothesize that VIP-induced proliferation in HT29 adenocarcinoma cells may involve a cAMP-Rap1/Ras-B-Raf-ERK signalling pathway.

Cell Division↗
PubMed1990

Suramin-induced differentiation of the human colic adenocarcinoma cell clone HT29-D4 in serum-free medium.

The clonal cell line HT29-D4 was able to grow in a completely defined medium containing EGF, selenous acid, and transferrin in the presence of the anti-helminthic drug suramin. In the absence of suramin, the kinetics of cell growth and the cell density obtained were dependent on the external EGF concentration. In the presence of suramin, cell density reached a plateau independent of EGF concentration above 50 ng/ml. At the morphological level, suramin allowed hemicyst formation in the cell monolayer. The cells were polarized with a well-ordered brush border facing the culture medium and mature junctional complexes that divided the cell membrane in two distinct domains. The carcinoembryonic antigen was found to be restricted to the apical membrane domain while the major histocompatibility molecules HLA-ABC were segregated within the basolateral domain. The electrical parameters of suramin-treated cells grown on permeable filters were measured and demonstrated that the cell monolayer was electrically active. These properties were never found in the absence of the drug. Moreover, the vasoactive intestinal polypeptide (VIP) was able to induce a dramatic increase in cAMP only when it was added, in agreement with the localization of the VIP receptor, in the lower compartment of the culture chamber. In conclusion we described for the first time conditions allowing the growth of functionally differentiated human colic cell monolayers in chemically defined medium. This model will contribute to a better understanding of suramin action and of the mechanisms involved in cell polarization.

Adenocarcinoma↗