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Colon Cancer Cells (colon + cancer_cell)
Kinds of Colon Cancer Cells Terms modified by Colon Cancer Cells Selected AbstractsE2F4 expression is required for cell cycle progression of normal intestinal crypt cells and colorectal cancer cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2009Hugo Garneau The generation of knock-out mice for E2F4 gene expression has suggested a role for this transcription factor in establishing and/or maintaining the intestinal crypt compartment. Having previously demonstrated that E2F4 is cytoplasmic in quiescent-differentiated cells but nuclear in growth factor-stimulated proliferative cells, the present study was aimed at determining the role of E2F4 in the control of human intestinal epithelial proliferation. Results herein demonstrate that lentiviral infection of an shRNA which specifically knocked-down E2F4 expression slowed down G1/S phase transition and the proliferation rate of normal human intestinal epithelial cells (HIEC) and of colon cancer cells. Protein expression of Cdk2, cyclins D1 and A, Cdc25A and c-myc was markedly down-regulated in shE2F4-expressing cells; by contrast, expression of the cell cycle inhibitors p21Cip/Waf and p27Kip1 was increased. In addition, the expression of many genes involved in DNA synthesis was down-regulated in shE2F4-expressing cells, whereas no modulation in E2F1 expression was observed. A decrease in E2F4 in colon cancer cell lines also resulted in a reduction in soft-agar growth capacity. Immunofluorescence experiments in human fetal intestine revealed that cells expressing high nuclear levels of E2F4 also expressed cyclin A protein. Lastly, E2F4 and its target cyclin A were up-regulated and mostly nuclear in human colorectal tumor cells in comparison to the corresponding benign epithelium. These results indicate that nuclear E2F4 may be determinant in the promotion of proliferation of human intestinal epithelial crypt cells and colorectal cancer cells. J. Cell. Physiol. 221: 350,358, 2009. © 2009 Wiley-Liss, Inc. [source] Mesalazine downregulates c-Myc in human colon cancer cells.ALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 12 2007A key to its chemopreventive action? Summary Background, Dysplasia and malignant transformation of colonocytes in ulcerative colitis are associated with overexpression of c-Myc and genes regulating cell survival. 5-Aminosalicylates such as mesalazine may reduce the development of colorectal cancer in ulcerative colitis, but the mechanisms of its chemopreventive action are not clear. Aims, To examine whether mesalazine affects the expression of c-Myc in human colon cancer cell lines. Methods, Human colon cancer cells were treated with vehicle or mesalazine (4 mm or 40 mm). We examined: (i) mRNA expression by gene array, (ii) protein expression by Western blotting and immunohistochemistry and (iii) apoptosis by Annexin V labelling. Results, Mesalazine significantly reduced expression of c-Myc mRNA and protein. Conclusions, Mesalazine downregulates gene and protein expression of c-Myc. The apoptotic and growth inhibitory effects of mesalazine are dose-dependent. Expression of c-Myc is significantly reduced by mesalazine 40 mm. [source] Synthesis and In-Vitro Antitumor Activities of Some Mannich Bases of 9-Alkyl-1,2,3,4-tetrahydrocarbazole-1-onesARCHIV DER PHARMAZIE, Issue 3 2009Jing Chen Abstract A novel series of 2-substituted aminomethyl-9-alkyl-1,2,3,4-tetrahydrocarbazole-1-ones 5a,q was synthesized via aminomethylation of 9-alkyl-1,2,3,4-tetrahydrocarbazole-1-ones 4a,e with hydrochlorides of the respective amines 6a,m. The structures of these newly synthesized compounds were characterized by 1H-NMR, MS, and elemental analysis. All the compounds were tested for their cytotoxic activity in vitro against four human tumor cell lines including human non-small lung cancer cells (A549), human gastric adenocarcinoma (SGC), human colon cancer cell (HCT116), human myeoloid leukemia cells (K562), and one multi-drug resistant subline (KB-VCR). Most compounds showed moderate to potent cytotoxic activity against the tested cell lines. Preliminary mechanism research indicated that the most promising compound, 2-diethylaminomethyl-9-methyl-1,2,3,4-tetrahydrocarbazole-1-one 5c, exhibited a potential inhibitory effect against microtubule. [source] Cover Picture: Electrophoresis 16'2010ELECTROPHORESIS, Issue 16 2010Article first published online: 19 AUG 2010 Issue no. 16 is a regular issue with an Emphasis on "Proteins and Proteomics" comprising 20 manuscripts distributed over 4 separate parts. Part I has 7 research articles on various aspects of proteins and proteomics including combinatorial peptide ligand library for accessing low abundance proteins, analysis of membrane proteins, proteomic profiling of human colon cancer cells, quantitative determinations of biomarkers in clinical diagnostics, recombinant factor VIII, analysis of E. coli soluble proteins, and a weakly basic amino-reactive fluorescent label for IEF of proteins and chip electrophoresis. Part II has 2 research articles dealing with the CE analysis of magnetic nanoparticles and a microfluidic magnetic bead impact for cell stimulation. Part III consists of 2 research articles dealing with on-line preconcentration in CE. Instrumentation, devices and various methodologies are described in 9 research articles, which make the content of Part IV. Featured articles include: Combinatorial peptide ligand library plasma treatment: Advantages for accessing low-abundance proteins ((doi: 10.1002/elps.201000188)) Precautions to improve the accuracy of quantitative determinations of biomarkers in clinical diagnostics ((doi: 10.1002/elps.201000243)) Rapid identification of Candida albicans in blood by combined capillary electrophoresis and fluorescence in situ hybridization ((doi: 10.1002/elps.201000138)) [source] Upregulation of glycolytic enzymes in proteins secreted from human colon cancer cells with 5-fluorouracil resistanceELECTROPHORESIS, Issue 12 2009Young-Kyoung Shin Abstract 5-Fluorouracil (5-FU) is the most commonly used chemotherapeutic agent for colorectal cancer (CRC). However, resistance to this drug is a major obstacle in CRC chemotherapy. Accurate prediction of response to 5-FU would avoid unnecessary chemotherapy and allow the selection of other effective drugs. To identify a candidate predictor of 5-FU resistance, we isolated secreted proteins that were up- or downregulated in a 5-FU-resistant cancer cell line, compared with the parent cell line (SNU-C4), using a stable isotope-coded labeling protocol. For validating the clinical applicability of this method, levels of the identified proteins were determined in the sera of 46 patients treated with 5-FU. In total, 238 proteins with molecular weights ranging from 50 to 75,kDa were identified. Among these, 45 and 35 secreted proteins were up- and downregulated in the 5-FU-resistant cell line, respectively. We observed significant upregulation of glycolytic enzymes, including glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase M2 (PK-M2), transketolase, and NADP(+)-dependent malic enzyme 1. In particular, the level of PK-M2, a key enzyme in the glycolytic pathway, showed an increasing tendency in both sera and tissues from CRC patients displaying no response to 5-FU-based chemotherapy (progressive and stable disease cases), compared with that in complete or partial responders to 5-FU-based chemotherapy; however, it did not reach the statistical significance. In conclusion, increasing pattern of PK-M2 observed with 5-FU resistance induced in vitro and in sera and tissues from CRC patients displaying poor response to 5-FU-based chemotherapy suggest the relevance of dysregulated glycolysis and 5-FU-resistant CRC. [source] NF-,B and apoptosis in colorectal tumourigenesisEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 5 2007M. M. Aranha Abstract Background, Nuclear factor-,B (NF-,B) may play an important role in colorectal tumourigenesis, controlling cell cycle and apoptosis gene expression. In addition, imbalances between cell proliferation and cell death are thought to underlie neoplastic development. The aims of this study were to investigate apoptosis and expression of several apoptosis-related proteins, and to determine correlations with colorectal tumour progression. Materials and methods, Apoptosis was evaluated by the TUNEL assay in 48 patient samples, including adenomas, adenocarcinomas and adjacent normal mucosas. Immunohistochemistry was performed for Bcl-2 and NF-,B. Expression levels of p53, Bax and I,B proteins were determined by immunoblotting. Cultured human colon cancer cells were used to evaluate NF-,B expression and nuclear translocation by immunocytochemistry and immunoblotting. Results, Apoptosis and NF-,B immunoreactivity were significantly higher in tumour tissue compared with normal mucosa (P < 0·01), increasing in association with histological tumour progression (P < 0·01). Bcl-2 was consistently higher in normal mucosa (P < 0·01) and inversely correlated with the percentage of apoptosis (P < 0·01). Phosphorylated p53 and Bax levels were similar in tumour tissue and normal mucosa; however, the NF-,B inhibitor, I,B, tended to decrease in tumours. In vitro, nuclear translocation of NF-,B was greater in proliferative than in resting phases of colon cancer cells. Conclusions, NF-,B expression and apoptosis are increased from adenoma to poorly differentiated adenocarcinoma tissues. Apoptosis is correlated with suppression of Bcl-2 expression, but appears to proceed through a p53- and Bax-independent pathway. Activation of NF-,B may play an important role in colorectal tumour progression. [source] COX-2-independent antiproliferative action of acetylsalicylic acid in human colon cancer cellsEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 11 2002M. N. Göke No abstract is available for this article. [source] The effects of acetylsalicylic acid on proliferation, apoptosis, and invasion of cyclooxygenase-2 negative colon cancer cellsEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 11 2002H.-G. Yu Summary Background Acetylsalicylic acid (ASA, aspirin), the most common nonsteroidal anti-inflammatory drug (NSAID), has been shown to have a protective effect against the incidence and mortality of colorectal cancer. However, the mechanism of its anticancer function remains unclear. The aim of this study was to determine the effects of acetylsalicylic acid on proliferation, apoptosis, and invasion in human cyclooxygenase-2 (COX-2) negative colorectal cancer cell lines. Materials and Methods After treatment with various concentrations of ASA, cell proliferation was measured in the human colon cancer cell line SW480. Apoptotic cells were identified by transmission electron microscopy, acridine orange staining, and flow cytometry. The invasive potential of SW480 cells was detected using an in vitro invasion assay. The production of carcinoembryonic antigen was measured by microparticle enzyme immunoassay. Expression of Bcl2, Bax, CD44v6, and nm23 were evaluated by immunocytochemistry. Results ASA significantly inhibited the proliferation of SW480 cells and stimulated apoptosis. Production of carcinoembryonic antigen and the invasive potential of SW480 cells were also inhibited by ASA. After treatment with ASA, down-regulation of Bcl2 and CD44v6 expression and up-regulation of nm23 expression were observed in SW480 cells. No obvious effect of ASA was found on Bax expression. Conclusion Our findings reveal that ASA inhibits the proliferation and promotes apoptosis in the human colon cancer cell line SW480. Down-regulation of Bcl2 expression might represent a potential mechanism by which ASA induces apoptosis in this COX-2 negative colon cancer cell line. Our results also suggest that ASA decreases the invasive potential of these colon cancer cells. Decreased CEA content and CD44v6 expression and elevated nm23 expression may contribute to the effect of ASA on invasive potential of SW480 colon cancer cells. [source] MicroRNA-143 reduces viability and increases sensitivity to 5-fluorouracil in HCT116 human colorectal cancer cellsFEBS JOURNAL, Issue 22 2009Pedro M. Borralho MicroRNAs are aberrantly expressed in cancer; microRNA-143 (miR-143) is down-regulated in colon cancer. HCT116 human colorectal cancer cells were used to investigate the biological role of miR-143. Transient miR-143 overexpression resulted in an approximate 60% reduction in cell viability. In addition, stable miR-143 overexpressing cells were selected with G418 and exposed to 5-fluorouracil. Increased stable expression of miR-143 was associated with decreased viability and increased cell death after exposure to 5-fluorouracil. These changes were associated with increased nuclear fragmentation and caspase -3, -8 and -9 activities. In addition, extracellular-regulated protein kinase 5, nuclear factor-,B and Bcl-2 protein expression was down-regulated by miR-143, and further reduced by exposure to 5-fluorouracil. In conclusion, miR-143 modulates the expression of key proteins involved in the regulation of cell proliferation, death and chemotherapy response. In addition, miR-143 increases the sensitivity of colon cancer cells to 5-fluorouracil, probably acting through extracellular-regulated protein kinase 5/nuclear factor-,B regulated pathways. Collectively, the data obtained in the present study suggest anti-proliferative, chemosensitizer and putative pro-apoptotic roles for miR-143 in colon cancer. [source] Intracellular trafficking and release of intact edible mushroom lectin from HT29 human colon cancer cellsFEBS JOURNAL, Issue 7 2000Lu-Gang Yu Our previous studies have shown that the Gal,1,3GalNAc,- (Thomsen,Friedenreich antigen)-binding lectin from the common edible mushroom Agaricus bisporus (ABL) reversibly inhibits cell proliferation, and this effect is a consequence of inhibition of nuclear localization sequence-dependent nuclear protein import after ABL internalization [Yu, L.G., Fernig, D.G., White, M.R.H., Spiller, D.G., Appleton, P., Evans, R.C., Grierson, I., Smith, J.A., Davies, H., Gerasimenko, O.V., Petersen, O.H., Milton, J.D. & Rhodes, J.M. (1999) J. Biol. Chem.274, 4890,4899]. Here, we have investigated further the intracellular trafficking and fate of ABL after internalization in HT29 human colon cancer cells. Internalization of 125I-ABL occurred within 30 min of the lectin being bound to the cell surface. Subcellular fractionation after pulse labelling of the cells with 125I-ABL for 2 h at 4 °C followed by culture of the cells at 37 °C demonstrated a steady increase in radioactivity in a crude nuclear extract. The radioactivity in this extract reached a maximum after 10 h and declined after 20 h. Release of ABL from the cell, after pulse labelling, was assessed using both fluorescein isothiocyanate-labelled ABL and 125I-ABL and was slow, with a t1/2 of 48 h. Most of the 125I-ABL both inside cells and in the medium remained intact, as determined by trichloroacetic acid precipitation and SDS/PAGE, and after 48 h only 22 ± 2% of ABL in the medium and 14 ± 2% inside the cells was degraded. This study suggests that the reversibility of the antiproliferative effect of ABL is associated with its release from cells after internalization. The internalization and subsequent slow release, with little degradation of ABL, reflects the tendency of lectins to resist biodegradation and implies that other endogenous or exogenous lectins may be processed in this way by intestinal epithelial cells. [source] The noncoding RNA, miR-126, suppresses the growth of neoplastic cells by targeting phosphatidylinositol 3-kinase signaling and is frequently lost in colon cancersGENES, CHROMOSOMES AND CANCER, Issue 11 2008Chunguang Guo MicroRNAs (miRNA/miR) are a class of small noncoding RNAs implicated in the pathogenesis of various malignancies. In the current study, using micro(RNA) arrays, we found a ubiquitous loss of miR-126 expression in colon cancer lines when compared to normal human colon epithelia. Reconstitution of miR-126 in colon cancer cells resulted in a significant growth reduction as evidenced in clonogenic assays. A search for miR-126 gene targets revealed p85,, a regulatory subunit involved in stabilizing and propagating the phosphatidylinositol 3-kinase (PI3K) signal, as one of the potential substrates. Restoration of miR-126 in cancer cells induced a ,3-fold reduction in p85, protein levels, with no concomitant change in p85,, a gene that is functionally related to p85, but not a supposed target of miR-126. Additionally, using reporter constructs, we show that the p85,-3, untranslated region is directly targeted by miR-126. Furthermore, this miR-126 mediated reduction of p85, was accompanied by a substantial reduction in phosphorylated AKT levels in the cancer cells, suggesting an impairment in PI3K signaling. Finally, in a panel of matched normal colon and primary colon tumors, each of the tumors demonstrated miR-126 down-regulation together with an increase in the p85, protein level. Taken together, we propose that miR-126 regulates PI3K signaling partly by targeting p85,, and that the loss of miR-126 may provide a selective growth advantage during colon carcinogenesis. © 2008 Wiley-Liss, Inc. [source] The dual EGFR/HER-2 tyrosine kinase inhibitor lapatinib sensitizes colon and gastric cancer cells to the irinotecan active metabolite SN-38INTERNATIONAL JOURNAL OF CANCER, Issue 12 2009Melissa J. LaBonte Abstract Members of the human epidermal receptor (HER) family are frequently associated with aggressive disease and poor prognosis in multiple malignancies. Lapatinib is a dual tyrosine kinase inhibitor targeting the epidermal growth factor receptor (EGFR) and HER-2. This study evaluated the therapeutic potential of lapatinib, alone and in combination with SN-38, the active metabolite of irinotecan (CPT-11), in colon and gastric cancer cell lines. Concentration-dependent antiproliferative effects of both lapatinib and SN-38 were observed in all colon and gastric cancer cell lines tested but varied significantly between individual cell lines (lapatinib range 0.08,11.7 ,M; SN-38 range 3.6,256 nM). Lapatinib potently inhibited the growth of a HER-2 overexpressing gastric cancer cell line and demonstrated moderate activity in gastric and colon cancer cells with detectable HER-2 expression. The combination of lapatinib and SN-38 interacted synergistically to inhibit cell proliferation in all colon and gastric cancer cell lines tested. Cotreatment with lapatinib and SN-38 also resulted in enhanced cell cycle arrest and the induction of apoptosis with subsequent cellular pharmacokinetic analysis demonstrating that lapatinib promoted the increased intracellular accumulation and retention of SN-38 when compared to SN-38 treatment alone. Finally, the combination of lapatinib and CPT-11 demonstrated synergistic antitumor efficacy in the LoVo colon cancer mouse xenograft model with no apparent increase in toxicity compared to CPT-11 monotherapy. These results provide compelling preclinical rationale indicating lapatinib to be a potentially efficacious chemotherapeutic combination partner for irinotecan in the treatment of gastrointestinal carcinomas. © 2009 UICC [source] Ascochlorin activates p53 in a manner distinct from DNA damaging agentsINTERNATIONAL JOURNAL OF CANCER, Issue 12 2009Ji-Hak Jeong Abstract Ascochlorin, a prenylphenol antitumor antibiotic, profoundly increases the expression of endogenous p53 by increasing protein stability in the human osteosarcoma cells and human colon cancer cells. Ascochlorin also increases DNA binding activity to the p53 consensus sequence in nuclear extract and enhances transcription of p53 downstream targets. Ascochlorin specifically induces p53 phosphorylation at ser 392 without affecting ser 15 or 20, whereas DNA damaging agents typically phosphorylate these serines. Moreover, ascochlorin does not induce phosphorylation of ATM and CHK1, an established substrate of ATR that is activated by genotoxins, nor does it increase DNA strand break, as confirmed by comet assay. The structure-activity relationship suggests that p53 activation by ascochlorin is related to inhibition of mitochondrial respiration, which is further supported by the observation that respiratory inhibitors activate p53 in a manner similar to ascochlorin. These results suggest that ascochlorin, through the inhibition of mitochondrial respiration, activates p53 through a mechanism distinct from genotoxins. © 2009 UICC [source] Adenovirus-mediated small hairpin RNA targeting Bcl-XL as therapy for colon cancerINTERNATIONAL JOURNAL OF CANCER, Issue 6 2007Hongbo Zhu Abstract Bcl-XL, an anti-apoptotic protein of Bcl-2 family, is overexpressed in colon cancers. To determine Bcl-XL's potential feasibility as a therapeutic target, we constructed a recombinant adenovirus that expressed a U6 promoter-driven small hairpin RNA (shRNA) targeting Bcl-XL (Ad/Bcl-XL shRNA) and evaluated the vector's ability to induce RNA interference in vivo and alter apoptosis induction in colon cancer cells and tumours. Ad/Bcl-XL shRNA effectively knocked down Bcl-XL expression in colon cancer cells and decreased their viability. Treatment with Ad/Bcl-XL shRNA but not control vectors led to dramatically increased cleavage of cellular apoptosis-related enzymes caspase-9, caspase-3 and poly(ADP-ribose) polymerase. Ad/Bcl-XL shRNA also significantly suppressed the growth of subcutaneous tumours derived from DLD1 cells in a nude mouse model and did so without causing any obvious damage to normal tissues or normal human fibroblasts. Together, our results support the feasibility of using adenovirus-mediated RNA interference therapy targeting Bcl-XL against colon cancers and warrant further studies of its safety and efficacy. © 2007 Wiley-Liss, Inc. [source] Disease-associated casein kinase I , mutation may promote adenomatous polyps formation via a Wnt/,-catenin independent mechanismINTERNATIONAL JOURNAL OF CANCER, Issue 5 2007I-Chun Tsai Abstract The Wnt signaling pathway is critical for embryonic development and is dysregulated in multiple cancers. Two closely related isoforms of casein kinase I (CKI, and ,) are positive regulators of this pathway. We speculated that mutations in the autoinhibitory domain of CKI,/, might upregulate CKI,/, activity and hence Wnt signaling and increase the risk of adenomatous polyps and colon cancer. Exons encoding the CKI, and CKI, regulatory domains were sequenced from DNA obtained from individuals with adenomatous polyps and a family history of colon cancer unaffected by familial adenomatous polyposis or hereditary nonpolyposis colorectal cancer (HNPCC). A CKI, missense mutation, changing a highly conserved residue, Arg324, to His (R324H), was found in an individual with large and multiple polyps diagnosed at a relatively young age. Two findings indicate that this mutation is biologically active. First, ectopic ventral expression of CKI,(R324H) in Xenopus embryos results in secondary axis formation with an additional distinctive phenotype (altered morphological movements) similar to that seen with unregulated CKI,. Second, CKI,(R324H) is more potent than wildtype CKI, in transformation of RKO colon cancer cells. Although the R324H mutation does not significantly change CKI, kinase activity in an in vitro kinase assay or Wnt/,-catenin signal transduction as assessed by a ,-catenin reporter assay, it alters morphogenetic movements via a ,-catenin-independent mechanism in early Xenopus development. This novel human CKI, mutation may alter the physiological role and enhance the transforming ability of CKI, through a Wnt/,-catenin independent mechanism and thereby influence colonic adenoma development. © 2006 Wiley-Liss, Inc. [source] Conjugated linoleic acid inhibits peritoneal metastasis in human gastrointestinal cancer cellsINTERNATIONAL JOURNAL OF CANCER, Issue 3 2006Hiroki Kuniyasu Abstract The effect of conjugated linoleic acid (CLA) on peritoneal metastasis was examined by in vitro treatment of cancer cells and mouse peritoneal metastasis models. First, cell growth of MKN28 human gastric cancer cells and Colo320 human colon cancer cells was suppressed by CLA in a dose-dependent manner with an increment in apoptosis. CLA significantly inhibited invasion into type IV collagen-coated membrane of MKN28 and Colo320 cells (p < 0.05). CLA-induced growth inhibition was recovered by the exposure to antisense S-oligodeoxynucleotide for peroxisome proliferator-activated receptor (PPAR)-, in both cell lines. BALB/c nu-nu mice were inoculated with MKN28 and Colo320 cells into their peritoneal cavity, and administrated with CLA intraperitoneally (weekly, 4 times). CLA treatment did not affect food intake or weight gain of mice. CLA treatment significantly decreased metastatic foci of both cells in the peritoneal cavity (p < 0.005). Survival rate in mice inoculated with MKN28 or Colo320 cells was significantly recovered by CLA treatment (p = 0.0025 and 0.0052, respectively). Protein production in MKN28 and Colo320 cells treated with CLA showed a decrease in epidermal growth factor receptor and transforming growth factor-, and an increase in Bax. These findings suggest that CLA inhibits metastasis of human gastric and colon cancer cells. © 2005 Wiley-Liss, Inc. [source] MRP1 and glucosylceramide are coordinately over expressed and enriched in rafts during multidrug resistance acquisition in colon cancer cellsINTERNATIONAL JOURNAL OF CANCER, Issue 4 2004Karin Klappe Abstract Previously we have described a novel multidrug-resistant cell line, HT29col, which displayed over expression of the multidrug-resistance protein 1 (MRP1) and an altered sphingolipid composition, including enhanced levels of glucosylceramide (GlcCer; Kok JW, Veldman RJ, Klappe K, Koning H, Filipeanu C, Muller M. Int J Cancer 2000;87:172,8). In our study, long-term screening revealed that, during colchicine-induced acquisition of multidrug resistance in a new HT29col cell line, increases in GlcCer occurred concomitantly with upregulation of MRP1 expression. Both MRP1 and GlcCer were found enriched in Lubrol-insoluble membrane domains. The expression of MRP1 and GlcCer were tightly correlated, as indicated also by a reversal of both at the later stage of colchicine consolidation. Resistance to colchicine was determined by MRP1, while glucosylceramide synthase (GCS) did not contribute: 1) Resistance was fully inhibited by MK571. 2) GCS expression and activity were not upregulated in HT29col cells. 3) Inhibition of GCS did not affect MRP1-mediated efflux function or sensitivity to colchicine. Instead, overall sphingolipid metabolism was upregulated through an increased rate of ceramide biosynthesis. In conclusion, upregulation of MRP1 occurs in concert with upregulation of GlcCer during multidrug-resistance acquisition, and both are enriched in rafts. The increased GlcCer pool does not directly modulate MRP1 function and cell survival. © 2004 Wiley-Liss, Inc. [source] 15-hydroxy-eicosatetraenoic acid arrests growth of colorectal cancer cells via a peroxisome proliferator-activated receptor gamma-dependent pathwayINTERNATIONAL JOURNAL OF CANCER, Issue 5 2003George G. Chen Abstract Peroxisome proliferator-activated receptor gamma (PPAR,) inhibits cell growth via promoting apoptosis. Human colorectal cancer tissues had abundant PPAR, but the incidence of apoptosis was very low, suggesting a defect in the PPAR, pathway. Here, we found that 15-hydroxy-eicosatetraenoic acid (15S-HETE), an endogenous ligand for PPAR,, was significantly decreased in the serum of patients with colorectal cancer. Treatment of colon cancer cells with 15S-HETE inhibited cell proliferation and induced apoptosis, which was preceded by an increase in TGF-,-inducible early gene (TIEG) and a decrease in Bcl-2. The action of 15S-HETE could be blocked when PPAR, was suppressed. Overexpression of Bcl-2 prevented the apoptosis. The levels of TIEG and 15-lipoxygenase (15-LOX), the enzyme responsible for 15S-HETE production, was decreased in colorectal cancer. Therefore, colorectal cancer is associated with decreased 15S-HETE. Treatment of colon cancer cells with 15S-HETE inhibits cell proliferation and induces apoptosis in a PPAR,-dependent pathway involving augmentation of TIEG and reduction of Bcl-2 expression. © 2003 Wiley-Liss, Inc. [source] Vitamin D Receptor: Key Roles in Bone Mineral Pathophysiology, Molecular Mechanism of Action, and Novel Nutritional Ligands,JOURNAL OF BONE AND MINERAL RESEARCH, Issue S2 2007Peter W Jurutka Abstract The vitamin D hormone, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], binds with high affinity to the nuclear vitamin D receptor (VDR), which recruits its retinoid X receptor (RXR) heterodimeric partner to recognize vitamin D responsive elements (VDREs) in target genes. 1,25(OH)2D3 is known primarily as a regulator of calcium, but it also controls phosphate (re)absorption at the intestine and kidney. Fibroblast growth factor 23 (FGF23) is a phosphaturic hormone produced in osteoblasts that, like PTH, lowers serum phosphate by inhibiting renal reabsorption through Npt2a/Npt2c. Real-time PCR and reporter gene transfection assays were used to probe VDR-mediated transcriptional control by 1,25(OH)2D3. Reporter gene and mammalian two-hybrid transfections, plus competitive receptor binding assays, were used to discover novel VDR ligands. 1,25(OH)2D3 induces FGF23 78-fold in osteoblasts, and because FGF23 in turn represses 1,25(OH)2D3 synthesis, a reciprocal relationship is established, with FGF23 indirectly curtailing 1,25(OH)2D3 -mediated intestinal absorption and counterbalancing renal reabsorption of phosphate, thereby reversing hyperphosphatemia and preventing ectopic calcification. Therefore, a 1,25(OH)2D3,FGF23 axis regulating phosphate is comparable in importance to the 1,25(OH)2D3,PTH axis that regulates calcium. 1,25(OH)2D3 also elicits regulation of LRP5, Runx2, PHEX, TRPV6, and Npt2c, all anabolic toward bone, and RANKL, which is catabolic. Regulation of mouse RANKL by 1,25(OH)2D3 supports a cloverleaf model, whereby VDR-RXR heterodimers bound to multiple VDREs are juxtapositioned through chromatin looping to form a supercomplex, potentially allowing simultaneous interactions with multiple co-modulators and chromatin remodeling enzymes. VDR also selectively binds certain ,3/,6 polyunsaturated fatty acids (PUFAs) with low affinity, leading to transcriptionally active VDR-RXR complexes. Moreover, the turmeric-derived polyphenol, curcumin, activates transcription of a VDRE reporter construct in human colon cancer cells. Activation of VDR by PUFAs and curcumin may elicit unique, 1,25(OH)2D3 -independent signaling pathways to orchestrate the bioeffects of these lipids in intestine, bone, skin/hair follicle, and other VDR-containing tissues. [source] HSP70 interacts with TRAF2 and differentially regulates TNF, signalling in human colon cancer cellsJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 3 2010Shengming Dai Abstract Members of tumour necrosis factor (TNF) family usually trigger both survival and apoptotic signals in various cell types. Heat shock proteins (HSPs) are conserved proteins implicated in protection of cells from stress stimuli. However, the mechanisms of HSPs in TNF,-induced signalling pathway have not been fully elucidated. We report here that HSP70 over-expression in human colon cancer cells can inhibit TNF,-induced NF,B activation but promote TNF,-induced activation of c-Jun N-terminal kinase (JNK) through interaction with TNF receptor (TNFR)-associated factor 2 (TRAF2). We provide evidence that HSP70 over-expression can sequester TRAF2 in detergent-soluble fractions possibly through interacting with TRAF2, leading to reduced recruitment of receptor-interacting protein (RIP1) and I,B, kinase (IKK) signalosome to the TNFR1,TRADD complex and inhibited NF,B activation after TNF, stimuli. In addition, we found that HSP70,TRAF2 interaction can promote TNF,-induced JNK activation. Therefore, our study suggests that HSP70 may differentially regulate TNF,-induced activation of NF,B and JNK through interaction with TRAF2, contributing to the pro-apoptotic roles of HSP70 in TNF,-induced apoptosis of human colon cancer cells. [source] ,-Catenin signaling in biological control and cancerJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2007Nancy Gavert Abstract A coordinated integration of cell,cell adhesion and the control of gene expression is essential for the development of multicellular, differentiated organisms. ,-Catenin fulfils important regulatory functions in both cell,cell adhesion by linking cadherin adhesion receptors to the cytoskeleton, and also as a key element in the Wnt signaling pathway where it acts as cotranscriptional activator of target genes in the cell nucleus. Wnt signaling is involved in numerous aspects of embryonic development and in the control of tissue self-renewal in a variety of adult tissues. Hyperactivation of Wnt signaling, mostly by affecting ,-catenin functions, is a hallmark of colon cancer and of many other human cancers. In this prospect, we discuss studies pointing to the molecular mechanisms that govern the integration between cell,cell adhesion and gene expression, as reflected in the switches between these two functions of ,-catenin in colon cancer cells. J. Cell. Biochem. 102: 820,828, 2007. © 2007 Wiley-Liss, Inc. [source] Methotrexate induced differentiation in colon cancer cells is primarily due to purine deprivationJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2006R. Singh Abstract The folate antagonist methotrexate (MTX) inhibits synthesis of tetrahydrofolate (THF), pyrimidines and purines, and induces differentiation in several cell types. At 1 µM, MTX reduced proliferation and induced differentiation in HT29 colon cancer cells; the latter effect was augmented (P,<,0.001) by thymidine (100 µM) but was reversed (P,<,0.001) by the purines, hypoxanthine (Hx; 100 µM) and adenosine (100 µM). In contrast 5-fluoro-uracil (5-FU), a specific thymidylate synthase (TS) inhibitor, had no effect on differentiation, suggesting that MTX-induced differentiation is not due to a reduction in thymidine but to the inhibition of purine biosynthesis. Inhibition of cyclic AMP (cAMP) by RpcAMP (25 µM) further enhanced (P,<,0.001) MTX induced differentiation, whereas the cAMP activator forskolin (10 µM) reversed (P,<,0.001) MTX induced differentiation. These observations implicate a central role of adenosine and cAMP in MTX induced differentiation. By combining Western blot analysis with liquid chromatography-mass spectrometry (LC-MS)and HPLC analyses we also reveal both the expression and activity of key enzymes (i.e. methionine synthase (MS), s-adenosylhomocysteinase, cystathionine ,-synthase and ornithine decarboxylase) regulating methyl cycle, transsulfuration and polyamine pathways in HT29 colon cancer cells. At 1 µM, MTX induced differentiation was associated with a marked reduction in the intracellular concentrations of adenosine and, consequently, S-adenosylmethionine (SAM), S-adenosylhomocysteine, polyamines and glutathione (GSH). Importantly, the marked reduction in methionine that accompanied MS inhibition following MTX treatment was non-limiting with respect to SAM synthesis. Collectively, these findings indicate that the effects of MTX on cellular differentiation and single carbon metabolism are primarily due to the intracellular depletion of purines. J. Cell. Biochem. © 2006 Wiley-Liss, Inc. [source] E2F4 expression is required for cell cycle progression of normal intestinal crypt cells and colorectal cancer cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2009Hugo Garneau The generation of knock-out mice for E2F4 gene expression has suggested a role for this transcription factor in establishing and/or maintaining the intestinal crypt compartment. Having previously demonstrated that E2F4 is cytoplasmic in quiescent-differentiated cells but nuclear in growth factor-stimulated proliferative cells, the present study was aimed at determining the role of E2F4 in the control of human intestinal epithelial proliferation. Results herein demonstrate that lentiviral infection of an shRNA which specifically knocked-down E2F4 expression slowed down G1/S phase transition and the proliferation rate of normal human intestinal epithelial cells (HIEC) and of colon cancer cells. Protein expression of Cdk2, cyclins D1 and A, Cdc25A and c-myc was markedly down-regulated in shE2F4-expressing cells; by contrast, expression of the cell cycle inhibitors p21Cip/Waf and p27Kip1 was increased. In addition, the expression of many genes involved in DNA synthesis was down-regulated in shE2F4-expressing cells, whereas no modulation in E2F1 expression was observed. A decrease in E2F4 in colon cancer cell lines also resulted in a reduction in soft-agar growth capacity. Immunofluorescence experiments in human fetal intestine revealed that cells expressing high nuclear levels of E2F4 also expressed cyclin A protein. Lastly, E2F4 and its target cyclin A were up-regulated and mostly nuclear in human colorectal tumor cells in comparison to the corresponding benign epithelium. These results indicate that nuclear E2F4 may be determinant in the promotion of proliferation of human intestinal epithelial crypt cells and colorectal cancer cells. J. Cell. Physiol. 221: 350,358, 2009. © 2009 Wiley-Liss, Inc. [source] Reduction of intracellular pH inhibits constitutive expression of Cyclooxygenase-2 in human colon cancer cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2004Daniela Pirkebner Cyclooxygenase-2 (COX-2) over-expression is critically involved in tumor formation. Intracellular pH (pHi) has been shown to be alkaline in cancer cells, and to be an important trigger for cell proliferation. This study therefore analyzed the relationship between pHi and COX-2 expression. HRT-18 and Caco-2 cells cultured in medium with bicarbonate maintained a pHi of ,7.6, which is higher than that of non-neoplastic cells. Cells grown in bicarbonate-free medium with a pH at 6.8 showed a reduction in pHi to approximately 7.0. Importantly, reduction of pHi resulted in a complete inhibition of COX-2 mRNA and protein expression. When cells were grown in bicarbonate-supplemented medium at pH 6.8, pHi maintained at ,7.6 and COX-2 expression was not inhibited. Additionally, analysis utilizing protein synthesis inhibitor cycloheximide demonstrated that pHi mediated inhibition of COX-2 mRNA expression requires de novo protein synthesis of regulatory protein(s). These data strongly suggest that an alkaline pHi is an important trigger for constitutive COX-2 expression. Defining pHi -mediated mechanisms that govern the constitutive COX-2 expression may help in developing new strategies to block COX-2 over-expression in cancer cells. J. Cell. Physiol. 198: 295,301, 2004© 2003 Wiley-Liss, Inc. [source] Epigenetics are involved in the regulation of the cell cycle and expression of tumor suppressor genes in human colon cancer cellsJOURNAL OF DIGESTIVE DISEASES, Issue 3 2003Ying Xuan CHEN OBJECTIVE: To investigate the effects of DNA methylation and histone acetylation on the cell cycle progression and expression of tumor suppressor genes in human colon cancer (HCC) cell lines. METHODS: Three HCC cell lines (HT-29, SW1116 and Colo-320) were treated with the DNA methylation inhibitor, 5-aza-2'-deoxycytidine (5-aza-dC) or/and histone deacetylase (HDAC) inhibitors, trichostatin A (TSA) or sodium butyrate. The methylation status of the promoter of the p16INK4A gene was assayed by methylation-specific PCR (MSP). The expression of p16INK4A and p21WAF1 was analyzed by RT-PCR. The cell cycle distribution was determined by flow cytometry. RESULTS: Before treatment, p16INK4A expression was slightly detected in the three cell lines (HT-29, SW1116 and Colo-320) and p21WAF1 expression was not detected in SW1116 and Colo-320 cells. The methylation level of the p16INK4A gene promoter significantly decreased and mRNA expression markedly increased in HT-29 cells after treatment with 1 µmol/L, but not 10 µmol/L, of 5-aza-dC for 24 h. In the SW1116 and Colo-320 cells, the expression of p16INK4A was markedly enhanced at 10 µmol/L or 5 µmol/L of 5-aza-dC for 24 h. However, p21WAF1 gene expression was not detected. Interestingly, after treatment with TSA or sodium butyrate, the transcription of p21WAF1 was significantly upregulated in these two cell lines. Furthermore, 5-aza-dC did not affect cell cycle distribution, but TSA or sodium butyrate blocked the cell cycle, mainly in the G1 phase. CONCLUSIONS: The expression of the p16INK4A gene is regulated by DNA methylation in three HCC cell lines. The expression of p21WAF1 gene is regulated by histone acetylation in SW1116 and Colo-320. In these two cell lines, histone hyperacetylation causes a G1 cell cycle arrest. [source] Aspirin and salicylate inhibit colon cancer medium- and VEGF-induced endothelial tube formation: correlation with suppression of cyclooxygenase-2 expressionJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 10 2003M. I. Shtivelband Summary., To determine whether aspirin and salicylate suppress colon cancer cell-mediated angiogenesis, we evaluated the effects of aspirin and sodium salicylate on endothelial tube formation on Matrigel. Aspirin and sodium salicylate concentration-dependently inhibited human endothelial cell (EC) tube formation induced by conditioned medium collected from DLD-1, HT-29 or HCT-116 colon cancer cells. Aspirin and sodium salicylate at pharmacological concentrations were equally effective in blocking tube formation. Neutralizing antivascular endothelial growth factor (VEGF) antibodies blocked colon cancer medium-induced tube formation. VEGF receptor 2 but not receptor 1 antibodies inhibited tube formation to a similar extent as anti-VEGF antibodies. These results indicate that VEGF interaction with VEGF receptor 2 is the primary mechanism underlying colon cancer-induced angiogenesis. Aspirin or sodium salicylate inhibited VEGF-induced tube formation in a concentration-dependent manner comparable to that of inhibition of colon cancer medium-induced endothelial tube formation. It has been shown that cyclooxygenase-2 (COX-2) is pivotal in cancer angiogenesis. We found that colon cancer medium-induced COX-2 protein expression in EC and aspirin or sodium salicylate suppressed the cancer-induced COX-2 protein levels at concentrations correlated with those that suppressed endothelial tube formation. Furthermore, aspirin and sodium salicylate inhibited COX-2 expression stimulated by VEGF. These findings indicate that aspirin and other salicylate drugs at pharmacological concentrations inhibit colon cancer-induced angiogenesis which is correlated with COX-2 suppression. [source] Mesalazine downregulates c-Myc in human colon cancer cells.ALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 12 2007A key to its chemopreventive action? Summary Background, Dysplasia and malignant transformation of colonocytes in ulcerative colitis are associated with overexpression of c-Myc and genes regulating cell survival. 5-Aminosalicylates such as mesalazine may reduce the development of colorectal cancer in ulcerative colitis, but the mechanisms of its chemopreventive action are not clear. Aims, To examine whether mesalazine affects the expression of c-Myc in human colon cancer cell lines. Methods, Human colon cancer cells were treated with vehicle or mesalazine (4 mm or 40 mm). We examined: (i) mRNA expression by gene array, (ii) protein expression by Western blotting and immunohistochemistry and (iii) apoptosis by Annexin V labelling. Results, Mesalazine significantly reduced expression of c-Myc mRNA and protein. Conclusions, Mesalazine downregulates gene and protein expression of c-Myc. The apoptotic and growth inhibitory effects of mesalazine are dose-dependent. Expression of c-Myc is significantly reduced by mesalazine 40 mm. [source] 1,1-bis(3,-indolyl)-1-(p -methoxyphenyl)methane activates Nur77-independent proapoptotic responses in colon cancer cellsMOLECULAR CARCINOGENESIS, Issue 4 2008Sung Dae Cho Abstract 1,1-Bis(3,-indolyl)-1-(p -methoxyphenyl)methane (DIM-C-pPhOCH3) is a methylene-substituted diindolylmethane (C-DIM) analog that activates the orphan receptor nerve growth factor-induced-B, (NGFI-B,, Nur77). RNA interference studies with small inhibitory RNA for Nur77 demonstrate that DIM-C-pPhOCH3 induces Nur77-dependent and -independent apoptosis, and this study has focused on delineating the Nur77-independent proapoptotic pathways induced by the C-DIM analog. DIM-C-pPhOCH3 induced caspase-dependent apoptosis in RKO colon cancer cells through decreased mitochondrial membrane potential which is accompanied by increased mitochondrial bax/bcl-2 ratios and release of cytochrome c into the cytosol. DIM-C-pPhOCH3 also induced phosphatidylinositol-3-kinase-dependent activation of early growth response gene-1 which, in turn, induced expression of the proapoptotic nonsteroidal anti-inflammatory drug-activated gene-1 (NAG1) in RKO and SW480 colon cancer cells. Moreover, DIM-C-pPhOCH3 also induced NAG-1 expression in colon tumors in athymic nude mice bearing RKO cells as xenografts. DIM-C-pPhOCH3 also activated the extrinsic apoptosis pathway through increased phosphorylation of c- jun N-terminal kinase which, in turn, activated C/EBP homologous transcription factor (CHOP) and death receptor 5 (DR5). Thus, the effectiveness of DIM-C-pPhOCH3 as a tumor growth inhibitor is through activation of Nur77-dependent and -independent pathways. © 2007 Wiley-Liss, Inc. [source] Differential apoptosis by gallotannin in human colon cancer cells with distinct p53 statusMOLECULAR CARCINOGENESIS, Issue 3 2007Sahar Al-Ayyoubi Abstract Gallotannin (GT), a plant polyphenol, has shown anticarcinogenic activities in several animal models including colon cancer. In our previous study, we showed that GT inhibits 1,2-dimethylhydrazine-induced colonic aberrant crypt foci and tumors in Balb/c mice, thus supporting a role for GT as a chemopreventive agent in colon cancer. However, at the molecular level, GT's mechanism of chemoprevention is still unclear. In this study, we aim at identifying GT's potential molecular mechanisms of action in in vitro studies. We show that GT differentially inhibits the growth of two isogenic HCT-116 (p53+/+, p53,/,) human colon cancer cells versus normal human intestinal epithelial cells (FHs 74Int). DNA flow cytometric analysis showed that GT induced S-phase arrest in both HCT-116 cell lines. Cell-cycle arrest in p53 (+/+) cells was associated with an increase in p53 protein levels and p21 transcript and protein levels. The inhibition of cell-cycle progression of HCT-116 p53 (+/+) cells by GT correlated with a reduction in the protein levels of cyclin D1, pRb, and the Bax/Bcl-2 ratio. Although GT did not induce apoptosis in p53 (+/+) cells, a significant induction of apoptosis was observed in p53 (,/,) cells as shown by TUNEL staining and flow cytometry analysis. Apoptosis induction in p53 (,/,) cells was associated with a significant increase in Bax/Bcl-2 protein levels. Our results demonstrate that GT inhibits the growth of HCT-116 colon cancer cells in a p53-independent manner but exhibits differential sensitivity to apoptosis induction in HCT-116 cells with distinct p53 status. © 2006 Wiley-Liss, Inc. [source] The cytotoxic effect of Bowman,Birk isoinhibitors, IBB1 and IBBD2, from soybean (Glycine max) on HT29 human colorectal cancer cells is related to their intrinsic ability to inhibit serine proteasesMOLECULAR NUTRITION & FOOD RESEARCH (FORMERLY NAHRUNG/FOOD), Issue 3 2010Alfonso Clemente Abstract Bowman,Birk inhibitors (BBI) from soybean and related proteins are naturally occurring protease inhibitors with potential health-promoting properties within the gastrointestinal tract. In this work, we have investigated the effects of soybean BBI proteins on HT29 colon adenocarcinoma cells, compared with non-malignant colonic fibroblast CCD-18Co cells. Two major soybean isoinhibitors, IBB1 and IBBD2, showing considerable amino acid sequence divergence within their inhibitory domains, were purified in order to examine their functional properties, including their individual effects on the proliferation of HT29 colon cancer cells. IBB1 inhibited both trypsin and chymotrypsin whereas IBBD2 inhibited trypsin only. Despite showing significant differences in their enzyme inhibitory properties, the median inhibitory concentration values determined for IBB1 and IBBD2 on HT29 cell growth were not significantly different (39.9±2.3 and 48.3±3.5,,M, respectively). The cell cycle distribution pattern of HT29 colon cancer cells was affected by BBI treatment in a dose-dependent manner, with cells becoming blocked in the G0,G1 phase. Chemically inactive soybean BBI had a weak but non-significant effect on the proliferation of HT29 cells. The anti-proliferative properties of BBI isoinhibitors from soybean reveal that both trypsin- and chymotrypsin-like proteases involved in carcinogenesis should be considered as potential targets of BBI-like proteins. [source] | ||||||||||||||||