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At least 865 records · Page 48Linked to original sources
PubMed2003

Additive interaction of oxaliplatin and 17-allylamino-17-demethoxygeldanamycin in colon cancer cell lines results from inhibition of nuclear factor kappaB signaling.

Elucidation of the mechanism by which oxaliplatin induces cell death is essential to enhancing its action. We investigated the effects of oxaliplatin and 17-allylamino-17-demethoxygeldanamycin (17-AAG) in a panel of four colon adenocarcinoma cell lines. Cytotoxicity assays demonstrated at least additivity in three of the cell lines. Activation of the c-Jun NH(2)-terminal kinase pathway by oxaliplatin does not determine cytotoxicity. Activation of p38 was shown to be a key proapoptotic mediator of oxaliplatin-induced cell death. Modulation of extracellular signal-regulated kinase and AKT signaling had no impact on oxaliplatin toxicity in these cells. Nuclear factor (NF)-kappaB was constitutively active in all of the cell lines and was inhibited by 17-AAG. Down-regulation of NF-kappaB transactivation by pharmacological inhibitors enhanced oxaliplatin cytotoxicity. These data support an interaction between 17-AAG and components of the NF-kappaB pathway in the modulation of oxaliplatin sensitivity in colon cancer cells.

Adenocarcinoma↗
PubMedDate not supplied

Relationship between differentiation-inducing activity and hypercalcemic activity of hexafluorotrihydroxyvitamin D3 derivatives.

Among 16 newly synthesized hexafluorotrihydroxyvitamin D3 derivatives, 24-Homo-26,26,26,27,27,27-hexafluoro (24H-F6)-1,24(S), 25(OH)3 vitamin D3 (VD3) (DD-011) [16] induced differentiation (i.e., appearance of NBT-positive cells) of human promyelocytic leukemic HL-60 cells most efficiently (EC50=0.5 nM), followed by 24H-F6-1,25(OH)2-22-oxa-VD3 (DD-006) [11] > F6-1,25(OH)2-VD3 (F6VD3) [2] > F6-1,25(OH)2-22-ene-VD3 (DD-009) [14] > 24H-F6-1,25(OH)2-VD3 (F6C28) [3] > 24H-F6-1,25(OH)2-1,23(S),25(OH)3-VD3 (DD-015) [18] > 24H-F6-1,25(OH)2-22-ene-VD3 (mvd1400) [6] > 22H-F6-1,25(OH)2-24-ene-VD3 (mvd3400) [5] > 24H-F6-1,23(R),25(OH)3-VD3 (DD-014) [17] > 24H-F6-1,22(S),25(OH)3-VD3 (DD-003) [7] > 24H-F6-1,22(S),25(OH)3-24-yne-VD3 (DD-005) [9] > 24H-F6-1,22(R),25(OH)3-24-yne-VD3 (mvd-1235) [10] > F6-1,25(OH)2-22-ene-VD3 (DD-008) [13] = 1,25(OH)2VD3 [1] (CC50=6 nM). On the other hand, 24H-F6-1,22(R),25(OH)3-VD3 (DD-004) [8], which is an isomer of DD-003 [7], showed much reduced activity (CC50=100 nM), suggesting the importance of the configuration of the OH group at the C-22. When their differentiation-inducing activity was plotted vs. the octanol-water partition coefficient (log P) used as a parameter of hydrophobicity, a bell-shaped curve was produced, with the bottom at log P=5.4-5.8. There was no clear-cut relationship between the differentiation-inducing activity and hypercalcemic activity (serum calcium elevating activity). Compounds [3, 7, 11, 17] showed relatively higher differentiation-inducing activity, with lesser hypercalcemic activity, as compared with [1]. Administration of [7] showed potent antiproliferation activity against colon cancer transplanted in nude mice. These results further confirmed the antitumor potential of hexafluorotrihydroxyvitamin D3 derivatives.

Animals↗
PubMed2005

[Change in apoptosis and its mechanism in intestinal epithelial cells under oxidative stress].

OBJECTIVE: To investigate change in apoptosis level and its mechanism of intestinal epithelial cells under oxidative stress. METHODS: HT-29 cells were cultured in vitro, which were treated with hydrogen peroxide (H2O2), to simulate the intestinal epithelial cells injured by reactive oxidative species. The cells viability was observed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, cell apoptosis and apoptosis associated proteins were evaluated by flow cytometry and Western blot. RESULTS: Cells viability of HT-29 was decreased by H2O2 which showed dose-dependent and time-dependent patterns (all P<0.05). The cell apoptotic ratios were increased with the concentration of H2O2 increased and the time of the stimulation prolonged compared with the controls (all P<0.05). Although the expression of the Bax was increased when HT-29 cells were stimulated with different concentrations of H2O2 for 24 hours, the expression of Bcl-2 was decreased. While HT-29 cells were stimulated with 500 micromol/L H2O2, the expression of the Bax was increased and that of Bcl-2 was decreased overtime. CONCLUSION: These data suggest that oxidative stress appears to be related to the apoptosis in intestinal epithelial cells under stress. The imbalance of Bcl-2/Bax expression might result in intestinal epithelial cell apoptosis in oxidative stress.

Apoptosis↗
PubMed2005

Do cancer cells selectively mutate HFE to increase their intracellular iron?

Malignant cells compared to their normal counter-parts, have an increased requirement for iron in order to achieve an enhanced cell growth. Transferrin receptor (TfR), an essential transport protein that enables cells to satisfy their need of iron, is upregulated in cancer cells. The hemochromatosis gene (HFE) produces a protein that interacts with TfR; and we hypothesized that tumor cells would selectively mutate HFE to improve their iron-uptake and thus provide themselves a growth advantage over non-tumor cells. A total of 36 non-small cell lung cancer (NSCLC) samples and matched tumor-free tissue from the same individuals were examined for HFE mutations using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). Sequencing analysis of the shifted bands in three of 36 cases (in both, the tumor and matching normal tissue of the same individuals) revealed the cysteine to tyrosine substitution at amino acid residue 282 (C282Y) in the protein. Surprisingly, matching samples from one patient with a heterozygous C282Y mutation in the germline, showed loss of heterozygosity in the tumor sample at the mutant HFE allele resulting in loss of the C282Y and retention of the normal allele. In addition, we examined 45 tumor cell lines from 11 different tissues. Five cell lines had a heterozygous mutation of HFE, none had a homozygous HFE mutation in the coding region. In summary, our experiments suggest that HFE mutations are not associated with a growth advantage for cancer cells.

Base Sequence↗
PubMed2006

A homologue of the Drosophila headcase protein is a novel tumor marker for early-stage colorectal cancer.

The altered expression of certain genes is frequently detected as a hallmark of malignant tumors. These differentially expressed genes have the potential to become molecular markers. We purified the fecal mRNA from patients with colorectal cancer to identify novel candidates by using oligo-nucleotide microarray hybridization. We identified 21 upregulated and 22 downregulated candidates with significantly altered expression in patient fecal samples that have not been previously characterized. A gene encoding a homologue of the Drosophila headcase protein (HECA) was further examined due to its high ranking and possible importance in some human cancers. A tendency towards increased expression of HECA was dependent upon the clinicopathological stage. In this report, healthy individuals expressed less HECA, either in their blood samples or feces. Moreover, we detected upregulated HECA in blood and fecal samples of patients with colorectal cancer, and its expression level was shown to be significantly correlated with disease status. Our data show that HECA may be an early-stage classifier of colorectal cancer that can discriminate between late- and early-stage disease. In conclusion, this study is the first to analyze differentially expressed genes in the feces of colorectal cancer patients using oligonucleotide microarrays. The data suggest that HECA expression levels in feces may be an effective classifier for early-stage colorectal cancer.

Biomarkers, Tumor↗
PubMed2006

Genetic analysis of peroxisome proliferator-activated receptor gamma1 splice variants in human colorectal cell lines.

Peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear hormone receptor family. In colon, this transcription factor is involved in differentiation of absorptive cells. PPARgamma participates also in colon carcinogenesis and cancer progression. Two isoforms, namely PPARgamma1 and PPARgamma2, have been described. Recently, new PPARgamma1 transcripts whose translation raises PPARgamma1 protein have been characterised. They differ from each other by combination of untranslated exons localised in the 5' UTR of the PPARG gene. Here, we studied whether such a diversity of PPARgamma transcripts occurs in human colon cell models. Based on bioinformatic analysis, putative untranslated exons were identified in the human PPARG gene. By RT-PCR analysis, we have demonstrated that several of these untranslated exons are included in PPARgamma transcripts from colon-derived cell lines or in those derived from other tissue. Using HT-29 cells, changes in PPARgamma1 mRNA levels were observed after treatment with PPARgamma agonists such as pioglitazone and troglitazone. These modifications correlated with particular PPARgamma transcripts excluding the untranslated exon A2. HT-29 cells treatment with actinomycin D or cycloheximide showed that the presence of PPARgamma mRNA including exon A2 was dependent on de novo protein synthesis. We concluded that diverse PPARgamma1 mRNA exist in colorectal cells. Levels of PPARgamma1 transcript varied according to the phenotype of colon cell model used. We suggest that regulation of PPARgamma1 mRNA levels could be dependent in part on the composition of untranslated exon(s) in the 5' UTR of PPARgamma1 mRNA.

Base Sequence↗
PubMedDate not supplied

Evaluation of the antitumor efficacy of the somatostatin structural derivative TT-232 on different tumor models.

The antitumor effects of the somatostatin structural derivative TT-232 in different rodent and xenograft tumor models are summarized in this report. TT-232 had previously been shown to inhibit the proliferation of a large number of cancer cell lines in vitro and reduce the size of different tumors in animal models in vivo. The effects of TT-232 by different routes of administration and treatment schedules were studied in various types of rodent and human xenograft tumor models. In the rodent tumor models S-180 sarcoma and P-388 lymphoid leukemia tumor the infusion treatment resulted in 76%-100% tumor growth inhibition and in 20%-60% of the mice being long-term and tumor-free survivors. In the aggressive C-26 colon carcinoma and MXT breast carcinoma, the TT-232 treatments resulted in 71%-75% tumor growth inhibition and an approximately 50% increased survival time. The tumor growth inhibitory effect of TT-232 on human tumor xenografts proved to be significant, resulting in 30%-80% decrease in tumor volume and in 20%-40% tumor-free animals. This antitumor efficacy of TT-232 was seen in almost all the tumors investigated. In our study, the route of infusion was shown to increase drug efficacy relative to conventional delivery methods. Our results suggested that TT-232 is an effective and promising antitumor agent.

Animals↗
PubMed2006

Molecular cloning and preliminary analysis of a fragile site associated gene.

OBJECTIVE: To analyze the molecular coining of a fragile site-associated gene. METHODS: Genomic Chinese hamster ovary (CHO) DNA library was constructed using high molecular weight CHO DNA partially digested with MboI restriction enzyme from cultured CHO cells. Screening of genomic DNA library followed the established procedures. Genomic CHO in the positive clones was sequenced. Appropriate primers were designed for the reverse transcriptase-polymerase chain reactions (RT-PCR). The RT-PCR products were cloned into a pCRII TOPO vector and confirmed by DNA sequencing. Antibodies were prepared using synthetic peptides as antigens by immunizing the rabbits. Immunohistochemical analyses were performed to evaluate the expression of the novel gene in different tissues. RESULTS: To investigate the molecular mechanism underlying the initial events of mdr1a amplification, we cloned 1q31 fragile site DNA. Strikingly, we found that this fragile site contained a novel gene which was designated as a fragile site-associated (FSA) gene. FSA encoded an unusually large mRNA of approximately16 kb. Full-length human FSA cDNA was cloned. FSA mRNA was expressed in many cultured cells and tissue types. Immunohistochemical analyses also revealed an expression pattern of the encoded proteins in postmitotic, well-differentiated epithelial compartments of many organs, including colon, mammary glands, ovary, prostate, and bladder. CONCLUSION: FSA plays an important role in regulating mammalian epithelial cell growth and differentiation.

Animals↗
PubMedDate not supplied

Dialysable factors from human leukocyte extracts activating human NK cytotoxicity against tumor cells: identification of factor-producing leukocyte subpopulations.

Dialysable human leukocyte extracts (10 kDa molecular weight cutoff) contain at least three cytotoxicity-stimulating factors CySF-L1, CySF-L2 and CySF-L3, activating natural killer (NK) cytotoxicity of human peripheral blood mononuclear cell (PBMC) against tumor cells when PBMC were pre-incubated with the factors for 72h prior to cytotoxicity assays. The factors could be separated by ion exchange chromatography on Dowex 50WX2. The leukocyte subpopulations producing these factors could be identified: CySF-L1 was detected in granulocyte extracts and T lymphocyte extracts, CySF-L2 in granulocyte extracts and monocyte extracts, CySF-L3 in T lymphocyte extracts and in B lymphocyte/NK cell extracts. All three factors were also present with low concentrations in fresh human plasma. The effector cells activated during pre-incubation of PBMC with the factors were partly identical: CySF-L1 and L2 activated the NK cytotoxicity of monocytes and of NK cells whereas CySF-L3 only stimulated the cytotoxicity of NK cells. All three factors stimulated LAK-like cytotoxicity since the effector cells showed enhanced activity not only against standard NK targets like K562 tumor cells but also against NK-insensitive targets like DAUDI cells and HT29 tumor cells.

Biological Factors↗
PubMed1988

Vasoactive intestinal peptide receptor/adenylate cyclase system: differences between agonist- and protein kinase C-mediated desensitization and further evidence for receptor internalization.

In this study we have characterized and compared the regulation of the HT29 cell vasoactive intestinal peptide receptor/adenylate cyclase system (VIP-R/AC) by the VIP-R agonist peptide histidineisoleucineamide (PHI) and by activators of protein kinase C (PKC) including phorbol 12-myristate, 13-acetate (PMA) and mezerein. Preincubation with either PHI or PKC activator decreased maximum VIP-stimulated AC activity and decreased the number of cell surface VIP-R. A [125I]VIP binding assay using solubilized VIP-R of the plasma membrane and light vesicle fractions from sucrose density step gradients was developed as a more direct measure of VIP-R internalization. Preincubation with PHI or PMA decreased plasma membrane fraction [125I]VIP binding and increased binding in the light vesicle fraction, thus providing the most direct evidence to date for translocation of VIP-R per se from the plasma membrane to another, presumably intracellular, compartment. Two experimental approaches differentiated between agonist and PKC activator regulation of VIP-R/AC. The protein kinase inhibitors H-7 and staurosporine blocked mezerein-, but not PHI-, induced losses of cell surface VIP-R. Also, down-regulation of PKC did not block PHI-induced loss of cell surface VIP-R. Thus, although both agonist and PKC activators can lead to desensitization and internalization of VIP-R, PKC is apparently not involved in the mechanisms of agonist-induced desensitization.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1996

Synthesis and anticancer properties of pyrimido[4,5-b]quinolines.

The synthesis and antitumor properties of pyrimido[4,5-b]quinolines have been described starting from 4-arylamino-5-carboxy-2-hydroxy-6-methylpyrimidines by reaction with PPA or POCl3. The reduced pyrimidoquinolines and derivatives with alkylating function have also been obtained.

Antineoplastic Agents↗
PubMed1996

Structure, functional expression, and cerebral localization of the levocabastine-sensitive neurotensin/neuromedin N receptor from mouse brain.

This work describes the cloning and expression of the levocabastine-sensitive neurotensin (NT) receptor from mouse brain. The receptor protein comprises 417 amino acids and bears the characteristics of G-protein-coupled receptors. This new NT receptor (NTR) type is 39% homologous to, but pharmacologically distinct from, the only other NTR cloned to date from the rat brain and the human HT29 cell line. When the receptor is expressed in Xenopus laevis oocytes, the H1 antihistaminic drug levocabastine, like NT and neuromedin N, triggers an inward current. The pharmacological properties of this receptor correspond to those of the low-affinity, levocabastine-sensitive NT binding site described initially in membranes prepared from rat and mouse brain. It is expressed maximally in the cerebellum, hippocampus, piriform cortex, and neocortex of adult mouse brain.

Amino Acid Sequence↗
PubMed1998

Bile acids inhibit tumour necrosis factor alpha-induced interleukin-8 production in human colon epithelial cells.

To clarify the regulatory mechanism of the production of various inflammatory mediators by intestinal epithelial cells, the effect of bile acids (tauroursodeoxycholate, TUDC; taurochenodeoxycholate, TCDC; and taurocholate, TC) on the cytokine-induced production of interleukin (IL)-8 in a human colon epithelial cell line (HT-29) was examined. HT-29 cells were incubated for 24 h in a culture medium containing tumour necrosis factor alpha (TNF alpha; 1 ng/mL) and/or interleukin (IL)-1beta (1 ng/mL) in the presence or absence of bile acids. The IL-8 concentration in the medium was measured by an enzyme-linked immunosorbent assay. The binding assay of TNF alpha was performed using [125I]-TNF alpha (100 pmol/L). Interleukin-8 production during incubation with TNF alpha was markedly reduced in the presence of 0.5 and 1 mmol/LTUDC, 0.5 and 1 mmol/LTCDC and 0.5 and 1 mmol/LTC, by 56, 85, 86, 91, 37 and 70%, respectively. The IL-8 production during incubation with IL-1beta was not significantly reduced in the presence of these bile acids. The specific binding of TNF alpha to cells was inhibited 33, 47, and 14% by 1 mmol/LTUDC, TCDC and TC, respectively. These findings suggest that bile acids inhibit TNF alpha-induced IL-8 production by the colonic cells. The suppression may be partly due to inhibition of TNF alpha binding to the cells by bile acids.

Bile Acids and Salts↗
PubMed1989

Comparison of vasoactive intestinal peptide-mediated protein phosphorylation in human lymphoblasts and colonic epithelial cells.

Vasoactive intestinal peptide (VIP) induces phosphorylation of a basic 38,000 mol. wt protein in a human lymphoblastic cell line (Molt 4b) and a human colon carcinoma cell line (HT29). In both cell types, VIP interacts with specific high affinity receptors to activate adenylate cyclase and cAMP-dependent protein kinase. The two cell types appear to express homologous receptors with similar affinity and specificity for VIP, but the colonic epithelial cells express a greater number of receptors. HT29 colonic cells also exhibit a greater stimulation of adenylate cyclase and a higher phosphorylation index for the 38,000 mol. wt protein in response to VIP. This 38,000 mol. wt protein, which is phosphorylated in the presence of VIP, appears to be identical in both cell lines; it is phosphorylated in both lymphoblasts and colonic epithelial cells in the presence of forskolin, but not in the presence of phorbol 12-myristate 13-acetate. Phosphorylation of this 38,000 mol. wt protein may be an important step in VIP regulation of water and electrolyte secretion from colonic epithelial cells, and in VIP regulation of immunoglobulin and lymphokine secretion from lymphocytes.

Adenylyl Cyclases↗
PubMed1998

Protein transport and processing by human HT29-19A intestinal cells: effect of interferon gamma.

BACKGROUND: The nature of the breakdown products produced in enterocytes during epithelial transport of intact proteins may be critical in determining the functional consequences of protein absorption. AIM: (a) To measure the transepithelial transport of horseradish peroxidase (HRP) and to identify the nature of HRP breakdown products released on the basal side of enterocytes and (b) to assess the role of interferon gamma (IFN gamma) on HRP transport and processing. METHODS: HT29-19A intestinal cells were used to assess transepithelial transport of HRP in Ussing chambers, and the nature of breakdown products in the basal compartment was analysed by high performance liquid chromatography (HPLC). RESULTS: (1) In control conditions, [3H]HRP equivalent fluxes (3135 (219) ng/h per cm2; mean (SEM) comprised 50% amino acids, 40% peptides, and 10% intact HRP. Steric exclusion HPLC of the breakdown products indicated a wide range of molecular masses including a major peptide of about 1150 Da. Lysosomal aspartyl and thiol proteases were expressed but no HLA-DR surface expression was noted, (2) At 48 to 72 hours after IFN gamma stimulation, [3H]HRP equivalent fluxes increased significantly (7392 (1433) ng/h per cm2) without modification of the relative proportions of amino acids, peptides, and intact HRP, and without modification of the distribution of breakdown products in HPLC. Lysosomal protease activities were not modified by IFN gamma but HLA-DR expression was increased. CONCLUSION: Intestinal cells are able to process HRP into peptides potentially capable of stimulating the immune system. IFN gamma stimulates the transport and processing of HRP thus increasing the antigenic load in the intestinal mucosa.

Amino Acids↗
PubMedDate not supplied

Recombinant human tumor necrosis factor and recombinant murine interleukin-1 alter the binding of Escherichia coli to intestine, mucin glycoprotein, and the HT29-C1 intestinal cell line.

Little is known about the effects of cytokines at the intestinal mucosal surface on the adherence of bacteria. We examined the effects of recombinant tumor necrosis factor (TNF) and interleukin-1 (IL-1) on the adherence of various strains of Escherichia coli to intestinal mucosa in vivo and in in vitro models. We studied the effects of TNF or IL-1 injected intraperitoneally on the ability of a rabbit enteric pathogen (RDEC-1) and a nonpathogenic E. coli (1392-) to colonize rabbit small bowel and found that there was a trend toward increased colonization by the RDEC-1 organisms in the TNF-treated rabbits, and a significant increase in colonization by the RDEC-1 organisms in the IL-1-treated animals (P < 0.01). Both TNF and IL-1 altered the density and the level of glycosylation of the small bowel mucus glcoprotein purified from the treated and untreated rabbits, and TNF treatment significantly increased the number of bacteria bound by this purified mucin (P < 0.01 for all strains). HT29-C1 intestinal cells in tissue culture were also grown in media with TNF or IL-1 and used in bacterial binding assays. The cells provided with media with 50 pg/mL of either cytokine bound significantly more of the three bacterial strains than cells in untreated media (P < 0.01 for all strains). The cytokines TNF and IL-1 have the potential to alter bacterial adherence to intestinal mucosa in vivo and in vitro; additional studies to clarify the role that these alterations in adherence may play in the clinical syndromes characterized by increased levels of intestinal cytokines are warranted.

Animals↗
PubMed2000

Interleukin-4 mediates cell growth inhibition through activation of Stat1.

Interleukin-4 (IL-4) activates Stat6 (signal transducer and activator of transcription 6) and plays multiple roles in regulation of the immune system. IL-4 also triggers phosphorylation of insulin receptor substrate (IRS), leading to stimulation of cell growth. Moreover, IL-4 inhibits proliferation of a variety of cells, but the molecular mechanism of its growth inhibitory effect is not understood. In this study, we demonstrated that IL-4 inhibited cell growth of colon carcinoma cell lines (HT29 and WiDr) but promoted cell growth of Burkitt's lymphoma cell lines (BL30 and BL41) in a dose-dependent manner. The growth inhibition was not dependent on Stat6 activation, because Stat6 was activated at similar levels in all cell lines in response to IL-4. Strikingly, IL-4 activated Stat1 in colon carcinoma cell lines but not in Burkitt's lymphoma cell lines. Therefore, these results suggest that IL-4 induced Stat1 activation, resulting in growth inhibition of colon carcinoma cell lines. Importantly, we present evidence that Stat1 is necessary for IL-4-mediated growth inhibition using Stat1-deficient and Stat1-reconstituted cells. The growth inhibitory effect of IL-4 was diminished in Stat1-deficient cells, whereas it was restored in Stat1-reconstituted cells. In addition, the expression of dominant-negative Stat1 in HT29 cells led to the loss of growth inhibition in response to IL-4. Taken together, our data suggest that IL-4 activates Stat1, leading to cell growth inhibition in colon cancer cells. Thus, this study demonstrates, for the first time, a molecular mechanism by which IL-4 inhibits cell growth.

Animals↗
PubMed1992

Identification of transglutaminase substrates in HT29 colon cancer cells: use of 5-(biotinamido)pentylamine as a transglutaminase-specific probe.

A biotinamine probe, 5-(biotinamido)pentylamine, was used for biotin-labeling of proteins in HT29 colon cancer cell extracts by endogenous transglutaminase activity. The biotin-labeled protein substrates were isolated and recovered by avidin-affinity chromatography. The proteins were separated using SDS-polyacrylamide gel electrophoresis, electroblotted onto a polyvinylidene difluoride membrane, visualized using Coomassie blue, cut out, and sequenced. Amino acid sequence data identified human fructose-1,6-bisphosphate aldolase A, an intracellular protein, as a substrate for cellular transglutaminase.

Amines↗