glycogen and Colonic-Neoplasms

Our previous studies showed that Candida tropicalis promoted colorectal cancer (CRC) by activating the function of MDSCs. However, underlying molecular mechanisms remains to be further investigated. In the present study, we indicated that C. tropicalis induced NLRP3 inflammasome activation through Dectin-3 in myeloid-derived suppressor cells (MDSCs). Mechanistically, we identified that C. tropicalis significantly enhanced the levels of glycolysis dependent on glycogen metabolism in MDSCs, which was required for NLRP3 inflammasome activation. C. tropicalis-induced NLRP3 inflammasome activation of MDSCs required the first priming signal and the second activation signal. For one thing, C. tropicalis promoted transcription of Nlrp3, Pro-caspase-1 and IL-1β genes through activation of JAK-STAT1 signaling pathway. For another, mtROS as the second activation signal mediated C. tropicalis-induced activation of NLRP3 inflammasome. Pharmacological inhibition of NLRP3 inflammasome activation abolished the pro-tumorigenic effect of C. tropicalis in an AOM/DSS-induced CAC mice model and significantly reduced C. tropicalis-promoted infiltration of MDSCs in colon tumors. Finally, in human CRC samples, the expression of STAT1, p-STAT1 and NLRP3 was elevated in MDSCs infiltrated by CRC. Collectively, these findings shed light on a previously unidentified mechanism by which C. tropicalis induces NLRP3 inflammasome activation in MDSCs to contribute to the progression of CRC. And STAT1-NLRP3 axis might represent a prospective therapeutic target for the treatment of CRC.

Topics: Animals; Candida tropicalis; Carcinogenesis; Colonic Neoplasms; Glycogen; Glycolysis; Humans; Inflammasomes; Mice; Myeloid-Derived Suppressor Cells; NLR Family, Pyrin Domain-Containing 3 Protein; Signal Transduction; STAT1 Transcription Factor

Topics: Amino Acid Transport System ASC; Cell Proliferation; Cell Transformation, Neoplastic; Colonic Neoplasms; Colorectal Neoplasms; Computational Biology; Exosomes; Fibroblasts; Fluorescent Antibody Technique; Glucose Transporter Type 1; Glycogen; Humans; Lactic Acid; Minor Histocompatibility Antigens; Proteomics; Tumor Microenvironment

Glycogen is the main energetic polymer of glucose in vertebrate animals and plays a crucial role in whole body metabolism as well as in cellular metabolism. Many methods to detect glycogen already exist but only a few are quantitative. We describe here a method using the Abcam Glycogen assay kit, which is based on specific degradation of glycogen to glucose by glucoamylase. Glucose is then specifically oxidized to a product that reacts with the OxiRed probe to produce fluorescence. Titration is accurate, sensitive and can be achieved on cell extracts or tissue sections. However, in contrast to other techniques, it does not give information about the distribution of glycogen in the cell. As an example of this technique, we describe here the titration of glycogen in two cell lines, Chinese hamster lung fibroblast CCL39 and human colon carcinoma LS174, incubated in normoxia (21% O2) versus hypoxia (1% O2). We hypothesized that hypoxia is a signal that prepares cells to synthesize and store glycogen in order to survive(1).

Topics: Animals; Cell Hypoxia; Cell Line; Cell Line, Tumor; Colonic Neoplasms; Cricetinae; Cricetulus; Fibroblasts; Fluorescence; Glucose; Glycogen; Humans; Hydrolysis; Oxidation-Reduction; Reagent Kits, Diagnostic; Titrimetry

The expression of the brush border-associated hydrolase sucrase-isomaltase was shown to increase from early to late passages of Caco-2 cells, concomitant with a decrease in the rates of glucose consumption. Twenty-six clones were isolated from early (P29) and late (P198) passages of the cell line. These clones show considerable and inverse differences in the levels of sucrase activities and rates of glucose consumption, without marked changes in other features of enterocytic differentiation of the cells (presence of an apical brush border, levels of expression of other brush border-associated hydrolases). Clones with low sucrase-isomaltase expression show a mosaic expression of the enzyme and a 38-fold higher rate of glucose consumption than clones with high sucrase-isomaltase expression. The clones with high expression show an homogeneous apical distribution of the enzyme and 70-fold and 35-fold higher levels of sucrase activities and sucrase-isomaltase mRNA, respectively. In contrast no differences were found from one clone to another in the enrichment of sucrase activity in brush border-enriched fractions as compared to cell homogenates. Switch to low glucose-containing medium (1 mM versus 25 mM in standard culture conditions) of cells with low sucrase-isomaltase results in an increased and more homogeneous expression of the enzyme and a tenfold augmentation of the levels of sucrase-isomaltase mRNA and sucrase activity. These results show that glucose interferes with the expression of sucrase-isomaltase in Caco-2 cells at the mRNA level.

Topics: Adenocarcinoma; Antibodies, Monoclonal; Cell Line; Clone Cells; Colonic Neoplasms; Fluorescent Antibody Technique; Gene Expression Regulation, Enzymologic; Glucose; Glycogen; Humans; Hydrolases; Kinetics; Microscopy, Electron; Microvilli; RNA, Messenger; Sucrase-Isomaltase Complex; Time Factors; Tumor Cells, Cultured

The potential use of Caco-2 cells as a model for the study of fructose metabolism and transport in the intestine was evaluated, since this human cell line exhibits many of the anatomical and biochemical characteristics of mature enterocytes. Pre- and postconfluent cultures converted [14C]fructose to CO2, lipid, and glycogen. Apparent utilization of [14C]fructose was less than that of [14C]glucose. This difference was due in part to the more rapid uptake of glucose from medium as compared with fructose. Addition of glucose, galactose, and mannose to medium markedly decreased the metabolism, while slightly inhibiting the uptake, of [14C]fructose. These data demonstrate that fructose can serve as a carbon and energy source for Caco-2 cells, and that common dietary monosaccharides affect the efficiency of fructose metabolism.

Topics: Adenocarcinoma; Biological Transport; Carbon Radioisotopes; Colonic Neoplasms; Epithelium; Fructose; Glucose; Glycogen; Humans; Kinetics; Lipid Metabolism; Monosaccharides; Radioisotope Dilution Technique; Tumor Cells, Cultured

Metabolic pathways of glucose utilization have been investigated in a human colon adenocarcinoma cell line (HT29) using carbon-13 Nuclear Magnetic Resonance spectroscopy. HT29 cells were adapted to grow on a polystyrene beaded microcarrier and were perfused when attached to the beads in a specially designed NMR cell. Abnormalities in carbohydrate metabolism already observed in several cancer cells were studied in HT29 cells fed with (1-13C)-enriched glucose. The cells were first perfused with a glucose-free medium for 2 h in order to deplete the intracellular store of glycogen, and they were subsequently perfused with a medium containing enriched glucose at an initial concentration of 5.5 mM. Sequential 13C-NMR spectra, recorded at 100.5 MHz (5 min accumulation), show that HT29 cells were able to utilize glucose through the glycolytic pathway while storing glucose as glycogen (glucose was utilized at a rate of 3.9 mumol/mg protein/hr). The glycolytic activity determined by the amount of lactic acid produced was 4.6 microns/mg protein/hr, corresponding to the formation of 1.2 lactic acid per glucose molecule. Glycogen accumulation corresponded to 16 micrograms/mg of protein. Treatment of HT29 with 10 nM vasoactive intestinal peptide (VIP) induced a transient decrease in the level of labelled glycogen to 50% of the initial value. Control level was recovered 12 min after VIP loading.

Topics: Adenocarcinoma; Cell Line; Colonic Neoplasms; Glycogen; Glycolysis; Humans; Lactates; Magnetic Resonance Spectroscopy; Time Factors; Tumor Cells, Cultured; Vasoactive Intestinal Peptide

Effects of insulin on key steps of carbohydrate metabolism were investigated in cultured HT29 colon cancer cells by two different approaches, i.e. incubation of the cells either in the absence or in the presence of glucose in the medium. In glucose-deprived cells, insulin decreased glycogen breakdown, but did not affect polysaccharide levels when glucose was present. Glycogen synthase became activated after insulin treatment in both conditions, even though the activation was more evident when glucose was omitted. No effect on glycogen phosphorylase activity was evident under our experimental conditions. In cells incubated with glucose, the hormone stimulated in a dose-dependent manner the rates of glucose uptake and lactate release. Concomitantly with the increase in glycolytic rate, insulin caused a strong increase in fructose 2,6-bisphosphate. This effect was not observed in the absence of glucose. It is concluded that the carbohydrate metabolism of cultured HT29 cells responds to insulin, making this biological model suitable for investigations in vitro on the mechanism of insulin action.

Topics: Adenocarcinoma; Colonic Neoplasms; Fructosediphosphates; Glucose; Glycogen; Glycogen Synthase; Glycolysis; Humans; Insulin; Lactates; Phosphorylases; Tumor Cells, Cultured

A previous report of members of our group demonstrated, in most adenocarcinomas and some adenomatous polyps with areas of malignant transformation, the presence of glycogen in cells that failed to show the synthesis of ordinary mucosubstances. The presence of glycogen in non-differentiated epithelium of the digestive tract of embryos, where it precedes ordinary mucus secretion, has lead some authors to suggest that the changes to glycogen secretion that occurs in malignant colonic cells, reflects the general regression to a less differentiated level of structure and function. It is well established that patients with carcinomas of the colon classified as less differentiated or higher grade tumors have significantly less chance of cure and consequently a worse prognosis than patients with lower grade tumors. The present ultrastructural study of the morphological differentiation characteristics of the glycogen containing cells previously encountered, revealed a strong association of glycogen to undifferentiated cells of the colonic adenocarcinomas and precancerous polypoid lesions studied. This undoubtedly seems to encourage the testing for the presence of glycogen in routine procedures as a possible marker of diagnostic and prognostic significance.

Topics: Adenocarcinoma; Adenoma; Colonic Neoplasms; Colonic Polyps; Glycogen; Histocytochemistry; Humans; Precancerous Conditions; Prognosis

Effect of glutamine deprivation (GLN- medium) and of its replacement by 4mM ammonium chloride (GLN-/NH4+ medium) or by 4mM glutamate (GLN-/Gt+ medium) was studied on growth rate, morphology and metabolism of HT29 human colon cancer cells. Growth rates were modified as follows: at the first passage, growth of GLN- cells was strongly decreased (doubling time: 192 hr vs 32 hr in control cells grown in GLN+ medium); GLN-/NH4+ cells and GLN-/Gt+ cells were found to have doubling times of 72 and 70 hr, respectively. At the 8th passage, doubling times were decreased in all cases, being: 144 hr for GLN- cells, 60 hr for GLN-/NH4+ cells and 24 hr for GLN-/Gt+ cells, which indicates a capacity of adaptation of the cell-line to new culture conditions. GLN- cells and GLN-/NH4+ cells were found to exhibit an enterocytic type of differentiation (polarization of the cell layer with apical and cystic brush border and tight junctions); GLN-/Gt+ cells remained undifferentiated and comparable to control GLN+ cells. Glycogen level varied according to the phases of the culture, with a trend to lower level in glutamine deprived cells; glucose uptake and lactate production varied as a function of the medium composition and of the phases of the culture. At the 8th passage, all the glutamine deprived cells produced less lactate than control; GLN-/Gt+ cells were found to utilize less glucose than others.

Topics: Ammonium Chloride; Cell Differentiation; Cell Division; Cell Line; Colonic Neoplasms; Glucose; Glutamates; Glutamic Acid; Glutamine; Glycogen; Humans; Lactates; Microscopy, Electron

Topics: Colon; Colonic Neoplasms; Esophagus; Female; Ganglioneuroma; Glycogen; Hamartoma; Humans; Hyperplasia; Middle Aged; Neoplasms, Multiple Primary; Syndrome

The human colon cancer line Caco-2 exhibits after confluency a concomitant increase of glycogen accumulation and an enterocytic differentiation. The purpose of this work was to investigate whether forskolin (FK), an activator of adenylate cyclase, would induce a permanent glycogenolysis and, if so, whether it would result in modifications of the differentiation pattern of the cells. FK activates adenylate cyclase in Caco-2 cells with an ED50 of 7 X 10(-6)M. Three different treatment protocols with FK (10(-5)M) were applied: 1) the cells were treated during all the time in culture (20 days); 2) the treatment was started after confluency; 3) the treatment was interrupted after confluency. The presence of FK results in a permanent stimulation of cAMP accumulation (10 to 20 fold the basal values) and in a permanently reduced glycogen content (30 or 50% of the control values). The rates of glucose consumption are increased three and five fold in protocols 1 and 3 respectively. These metabolic changes are associated with morphological changes (tightening of the intercellular spaces and shortening of the brush border microvilli) and with a dual inhibition of the activities of brush border hydrolases: a) an inhibition of the post-confluent increase of activity of sucrase, aminopeptidase N and alkaline phosphatase in the brush border enriched fraction; b) an inhibition of the post-confluent increase of activity of sucrase in the cell homogenate. A comparison of the results obtained in each protocol shows that the morphological modifications and the decrease of the enzyme activities in the brush border fraction are regularly associated with an increased cAMP accumulation, whereas the inhibition of the differentiation of sucrase is a direct consequence of the increase in glucose consumption and decrease in glycogen stores.

Topics: Cell Line; Cell Transformation, Neoplastic; Colforsin; Colonic Neoplasms; Cyclic AMP; Diterpenes; Dose-Response Relationship, Drug; Glucose; Glycogen; Humans; Hydrolases; Lactates; Microvilli

The activities of glycogen synthase and phosphorylase were measured and compared to the growth-related variations of glycogen accumulation in three cultured human tumor cell lines: HT-29 (colon carcinoma); MeWo (malignant melanoma); and RT-4 (carcinoma of the urinary bladder). A similar pattern of variations in the enzyme activities was found in the three cell lines. The activities of the a + b forms of glycogen phosphorylase increased throughout the culture period. Maximal activity of phosphorylase a coincided with low intracellular concentrations of glycogen during the period of exponential growth. When the rate of cell division decreased, phosphorylase a activity also decreased while the glycogen levels increased. Glycogen synthase was almost entirely in b form during the entire culture period, i.e., in both the exponential and the stationary phases. In vitro incubation of the cellular extracts without NaF showed, however, that the enzyme could be partially converted to the a form by the endogenous phosphatases. The A0.5 values of the enzyme for glucose-6-phosphate (Glc-6-P) were of the same order of magnitude as the intracellular Glc-6-P concentrations which ranged from 2.2 to 5.4 mM (almost 10 times those reported in normal cells). Similar Glc-6-P values were obtained by two different extraction methods controlled by the intracellular ATP and ADP concentrations. The Km values for uridine-5'-diphosphoglucose were always 2 to 3 times lower than the intracellular uridine-5'-diphosphoglucose concentrations. These results suggest that: (a) in these tumor cells, glycogen is essentially synthesized by glycogen synthase b via an allosteric activation by intracellular Glc-6-P; (b) there is no obvious growth-related control of glycogen synthase activity; and (c) the activity of glycogen phosphorylase seems to be growth dependent with maximal phosphorylase a activities associated with the period of high division rate.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Cell Line; Colonic Neoplasms; Glucose-6-Phosphate; Glucosephosphates; Glycogen; Glycogen Synthase; Humans; Kinetics; Melanoma; Neoplasms; Uridine Diphosphate Glucose; Urinary Bladder Neoplasms

The neutrophil glycogen content, as revealed by cytochemical staining, was determined in 22 patients with gastric cancer, 8 patients with cancer of large intestine and 40 healthy subjects. A statistically significant increase of the glycogen content in neutrophils obtained from patients with initial cancer stages as compared with healthy individuals and patients with advanced tumors was observed. The possible mechanism of these changes is discussed.

Topics: Colonic Neoplasms; Glycogen; Humans; Neutrophils; Stomach Neoplasms

Activity of several enzymes of the glycogen and carbohydrate metabolism is studied in HT 29 colon adenocarcinoma cell line and in HT 29 tumors developed in nude mice, by reference to the normal human colon mucosa. Activity of glycogen synthase, glycogen phosphorylase, pyruvate kinase, fructose-1,6-diphosphatase, glucose-6-phosphate dehydrogenase and lactate dehydrogenase is found to be increased in both the cultured cells and the tumors. It indicates that the biochemical strategy of malignant cells, due to the neoplastic transformation process, involves specific changes in the carbohydrate metabolism of tumor as well as in vitro growing correspondent cell line.

Topics: Adenocarcinoma; Animals; Carbohydrate Metabolism; Cell Line; Colonic Neoplasms; Enzymes; Glucosephosphate Dehydrogenase; Glycogen; Mice; Mice, Nude; Pyruvate Kinase

The effect of sodium butyrate on three cultured human colon carcinoma cell lines was studied. Exposure to butyrate caused morphological changes and resulted in the alteration of several growth properties. Doubling times of treated cells were increased five-fold and saturation densities and cloning efficiencies were decreased, compared to untreated cells. Histochemical studies using the periodic acid-Schiff reaction in conjunction with diastase digestion showed that butyrate induced increased glycogen levels in all three cell lines. This increase was confirmed by biochemical techniques. These effects of butyrate were reversed when treated cells were subsequently grown in the absence of butyrate. These changes are consistent with findings from several laboratories that butyrate can induce phenotypic changes in cultured tumour cells.

Topics: Butyrates; Butyric Acid; Cell Line; Clone Cells; Colonic Neoplasms; Glycogen; Humans

The serologically active moiety of an antigen detected occasionally in pathologic sera by double-diffusion gel precipitation tests, referred to in earlier studies as ubiquitous tissue antigen, was identified as a dextran composed predominantly, if not exclusively, of a(1 leads to 6)-linked glycopyranoses. By means of an enzyme immunoassay, dextran or dextran-like material, which inhibited the binding of antidextran serum to dextran, was detected in sera of several patients with various gastrointestinal (GI) diseases, especially GI ulcers (7/10), and also often in sera of aged people (9/21). However, 2 of 50 normal blood donor sera and a few sera from almost every disease group studied contained low quantities of dextran-like material. The levels of antidextran antibodies of the IgG class were also often elevated among patients with GI diseases and aged people as demonstrated by enzyme immunoassay with dextran T-500 as the solid phase antigen. OD values exceeding the mean plus 2.5 SD of 106 normal blood donor sera were recorded in ;69% of patients with gastric and duodenal ulcers, 40% with ulcerative colitis, 29% with Crohn's disease, 20% with colorectal carcinomas, and in 21% with rheumatoid arthritis. None of 23 children, but 9 of 23 aged people (35%) had elevated antibody levels. It is suggested that absorption of dextrans from food or their production by intestinal bacteria may be facilitated in various GI diseases.

Topics: Adult; Aged; Animals; Antibody Formation; Cattle; Chemical Precipitation; Child; Colonic Neoplasms; Dextrans; Diverticulitis; Epitopes; Glycogen; Humans; Immunodiffusion; Immunoenzyme Techniques; Kidney Diseases; Ostreidae; Rabbits

Topics: Adenocarcinoma; Cell Line; Colonic Neoplasms; Cyclic AMP; Gastrointestinal Hormones; Glycogen; Humans; In Vitro Techniques; Phosphorylase a; Vasoactive Intestinal Peptide

Topics: Adenocarcinoma; alpha-Glucosidases; Animals; Cell Line; Colonic Neoplasms; Disaccharides; Glucosidases; Glucosides; Glycogen; Humans; Hymecromone; Kinetics; Maltose; Mice; Mice, Nude

Of a total of 544 tubular, villous and tubulo-villous adenomas of the human colon which were investigated by light microscopy, six tubular and six villous adenomas were examined under the electron microscope. It was shown that the two types of adenoma differ in their tissue architecture, but not in their cytological appearance. Different grades of epithelial atypia occur in both types of adenoma. These are designated as grades I to III, correspond to mild, moderate and severe atypia respectively. Whereas adenoma cells with atypia grade I clearly show a cytological relationship with crypt epithelia of the normal colonic mucosa under the electron microscope, adenoma cells with atypia grade III have largely lost the differentiation characteristics of the parent cells and there is no longer intracytoplasmic organization. In the cytoplasm of adenoma cells with atypia grade II, glycogen storage is found as a special feature which does not occur in normal colonic epithelium. A similar finding has been described in the course of malignant transformation of the liver and kidney and thus can be regarded as a further indication of the precancerous nature of the adenomas.

Topics: Adenoma; Cell Differentiation; Colonic Neoplasms; Glycogen; Humans; Microscopy, Electron

Topics: Carcinoma; Cell Cycle; Cell Line; Colonic Neoplasms; Glycogen; Humans

Topics: Adenocarcinoma; Animals; Colon; Colonic Neoplasms; Epithelium; Glycogen; Humans; Intestinal Mucosa; Mice

The presence and kinetics of intracellular glycogen levels were studied, in relationship to cell growth, in asynchronous and in synchronized cultures of four human colorectal adenocarcinoma cell lines (HT-29, HRT-18, SW-480, and Caco-2). The results show that a specific pattern of glycogen accumulation occurs during the process of cell growth of the studied cell lines. The kinetics of glycogen accumulation in asynchronous cultures were similar from one cell line to another and were characterized by a low amount in the exponential phase of growth, followed by a 3- to 4-fold increase in the stationary phase. The quantities found in either phase were specific for each cell line. The maximum values found in Caco-2, HRT-18, HT-29, and SW-480 cells were, respectively, 258.5 +/- 6.9 (S.D.), 88.9 +/- 2.6, 87.5 +/- 3, and 17.5 +/- 1.8 microgram of glycogen per mg of proteins. The kinetics of glycogen accumulation during the cell cycle was also studied in synchronized cultures of HT-29 and HRT-18 cell lines. Both cell lines exhibited a common pattern of low glycogen quantities during S, G2, and M followed by an increase beginning with G1 and peaking (2.5 to 3 times the initial values) in the middle of this phase. This was followed by a symmetrical decrease in the second half of G1.

Topics: Adenocarcinoma; Cell Cycle; Cell Line; Colonic Neoplasms; Glycogen; Kinetics; Neoplasms, Experimental; Rectal Neoplasms

Rabbit antisera raised against a strain of E. coli 013, with a strong antiglycogen activity, were tested on human fetal and normal adult colons, on colon carcinomas, and on colon tumor cells in culture (HT29). Only very rare granules were present in adult normal colons when tested with the immunofluorescence method. In faetal colons, in 12 out of 14 carcinomas, and on HT29 cells, the immunofluorescent reactions were similar to those observed in normal liver. The reactions were negative after previous treatment with alpha-amylase. They were inhibited with glycogen, with phenol-alcohol, perchloric, and trichloroacetic extracts from faetal colons, and with a tumor trichloroacetic extract. The extracts precipitated with anti-E. coli 013 antisera. They had a strong inhibiting activity in a radioimmunoassay test with labeled glycogen. The extracts from normal adult colons did not precipitate with the antisera and they had no inhibiting activity in either immunofluorescence and radioimmunoassay tests.

Topics: Adult; Antibodies, Bacterial; Carcinoma; Colon; Colonic Neoplasms; Escherichia coli; Female; Glycogen; Humans; Immune Sera; Neoplasms, Experimental; Pregnancy

Topics: Acetates; Chondroitin; Colon; Colonic Neoplasms; Glycogen; Histocytochemistry; Humans; Hyaluronic Acid; Intestinal Mucosa; Intestinal Polyps; Keto Acids; Neuraminic Acids; Papilloma; Periodic Acid; Polysaccharides; Staining and Labeling; Sulfates

Topics: Adenocarcinoma; Colon; Colonic Neoplasms; Glycogen; Glycosaminoglycans; Histocytochemistry; Humans; Hyaluronic Acid; Keto Acids; Neuraminic Acids; Periodic Acid; Plasma Cells; Polysaccharides; Staining and Labeling; Sulfates

Topics: Colonic Neoplasms; Glycogen; Humans