Cancerous Intestinal Cells Research Articles

Stem Cell Properties of Gastric Cancer Stem-Like Cells under Stress Conditions Are Regulated via the c-Fos/UCH-L3/β-Catenin Axis.

Gastric cancer stem-like cells (GCSCs) possess stem cell properties, such as self-renewal and tumorigenicity, which are known to induce high chemoresistance and metastasis. These characteristics of GCSCs are further enhanced by autophagy, worsening the prognosis of patients. Currently, the mechanisms involved in the induction of stemness in GCSCs during autophagy remain unclear. In this study, we compared the cellular responses of GCSCs with those of gastric cancer intestinal cells (GCICs) whose stemness is not induced by autophagy. In response to glucose starvation, the levels of β-catenin and stemness-related genes were upregulated in GCSCs, while the levels of β-catenin declined in GCICs. The pattern of deubiquitinase ubiquitin C-terminal hydrolase-L3 (UCH-L3) expression in GCSCs and GCICs was similar to that of β-catenin expression depending on glucose deprivation. We also observed that inhibition of UCH-L3 activity reduced β-catenin protein levels. The interaction between UCH-L3 and β-catenin proteins was confirmed, and it reduced the ubiquitination of β-catenin. Our results suggest that UCH-L3 induces the stabilization of β-catenin, which is required to promote stemness during autophagy activation. Also, UCH-L3 expression was regulated by c-Fos, and the levels of c-Fos increased in response to autophagy activation. In summary, our findings suggest that the inhibition of UCH-L3 during nutrient deprivation could suppress stress resistance of GCSCs and increase the survival rates of gastric cancer patients.

Oral administration of M13-loaded nanoliposomes is safe and effective to treat colitis-associated cancer in mice

ABSTRACT Objective Colitis-associated cancer (CAC) treatment lacks effective small-molecule drugs and efficient targeted delivery systems. Here, we loaded M13 (an anti-cancer drug candidate) to colon-targeting ginger-derived nanoliposomes (NL) and investigated if orally administered M13-NL could enhance the anticancer effects of M13 in CAC mouse models. Methods The biopharmaceutical properties of M13 were assessed by physicochemical characterizations. The in vitro immunotoxicity of M13 was assessed against PBMCs using FACS and the mutagenic potential of M13 was evaluated by the Ames assay. The in vitro efficacy of M13 was tested in 2D- and 3D-cultured cancerous intestinal cells. AOM/DSS-induced CAC mice were used to evaluate the therapeutic effects of free M13 or M13-NL on CAC in vivo. Results M13 has beneficial physiochemical properties, including high stability, and no apparent immunotoxicity or mutagenic potential in vitro. M13 is effective against the growth of 2D- and 3D-cultured cancerous intestinal cells in vitro. The in vivo safety and efficacy of M13 were significantly improved by using NL for drug delivery (p < 0.001). Oral administration of M13-NL exhibited excellent therapeutic effects in AOM/DSS-induced CAC mice. Conclusion M13-NL is a promising oral drug formulation for CAC treatment.

ORAL ADMINISTRATION OF NANOLIPOSOMES LOADED M13 MAY TREAT ULCERATIVE COLITIS AND COLITIS-ASSOCIATED CANCER WITHOUT CAUSING ANY SIDE EFFECTS

Abstract Intestinal inflammation, especially chronic (such as inflammatory bowel disease [IBD]), is an indispensable participant in the colonic neoplastic process, fostering the proliferation, survival, and migration of colonic tumor cells. Molecularly targeted therapeutics (e.g. anti-TNF-α agents) that exploit the targets involved in the pathophysiology of inflammatory responses in IBD are the most potent drugs currently available to treat IBD, which may also present anti-colon cancer effects. Unfortunately, most of these drugs were administered systemically in a high dose, and their use is limited by severe side effects. There is an unmet need for a targeted carrier system capable of delivering drugs specifically and effectively to inflamed colonic regions over a prolonged period. Such a system could significantly reduce drug dosage and side effects that limit existing treatments. The goal of the present study is to engineer natural “nanoliposomes” from the lipids of ginger-derived-nanoparticles and load with an anti-inflammatory drug candidate (M13), test its anticancer activity, and evaluate its safety. MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay demonstrated that M13, a stable metabolite from 6-Shogaol as demonstrated by the incubation with intestinal microsomes, was more cytotoxic against cancerous intestinal cells (e.g. HCT-116, Caco-2/BBe, and human colon cancer stem cell) than normal intestinal cells (e.g. HIEC-6, FHC, and human colon stem cell). The results suggested that M13 has more a better ability to specifically target cancerous intestinal cells over normal intestinal cells. Protein point hybridization data showed that nanoliposomes-loaded M13 decreased the expression levels of proteins involved in colon cancer progressions such as Survivin, EGFR, and BCL-x in colon cancer cells. Further, orally administered nanoliposomes-loaded M13 presented potent wound-healing/anti-inflammatory effects in the DSS-induced mouse model of colitis and anti-cancer effects in the AOM/DSS-induced colon cancer model. We also demonstrated that nanoliposomes-loaded M13 treatment did not affect blood. In addition, we examined the anti-inflammatory activity of M13 on active ulcerative mucosal biopsies (Reprocell tissue protocol TPS-008) from a patient (42-year-old white male) undergoing therapeutic resection for ulcerative colitis. The results of the RT2 Profiler PCR Array demonstrated that M13 had very potent anti-inflammatory activity and reduced the expression levels of the pro-inflammatory cytokines involved in IBD, including TNF-α, IL-1β, IL-6, and IL-23A, in the M13-treated human ulcerative biopsies compared to the untreated controls. Our proposed targeted therapy using nanoliposomes that deliver M13 to the colon might be used to treat ulcerative colitis and colitis-associated cancer without causing any side effects.

ORAL ADMINISTRATION OF NANOLIPOSOMES LOADED M13 MAY TREAT ULCERATIVE COLITIS AND COLITIS-ASSOCIATED CANCER WITHOUT CAUSING ANY SIDE EFFECTS

Intestinal inflammation, especially chronic (such as inflammatory bowel disease [IBD]), is an indispensable participant in the colonic neoplastic process, fostering the proliferation, survival, and migration of colonic tumor cells. Molecularly targeted therapeutics (e.g. anti-TNF-α agents) that exploit the targets involved in the pathophysiology of inflammatory responses in IBD are the most potent drugs currently available to treat IBD, which may also present anti-colon cancer effects. Unfortunately, most of these drugs were administered systemically in a high dose, and their use is limited by severe side effects. There is an unmet need for a targeted carrier system capable of delivering drugs specifically and effectively to inflamed colonic regions over a prolonged period. Such a system could significantly reduce drug dosage and side effects that limit existing treatments. The goal of the present study is to engineer natural “nanoliposomes” from the lipids of ginger-derived-nanoparticles and load with an anti-inflammatory drug candidate (M13), test its anticancer activity, and evaluate its safety. MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay demonstrated that M13, a stable metabolite from 6-Shogaol as demonstrated by the incubation with intestinal microsomes, was more cytotoxic against cancerous intestinal cells (e.g. HCT-116, Caco-2/BBe, and human colon cancer stem cell) than normal intestinal cells (e.g. HIEC-6, FHC, and human colon stem cell). The results suggested that M13 has more a better ability to specifically target cancerous intestinal cells over normal intestinal cells. Protein point hybridization data showed that nanoliposomes-loaded M13 decreased the expression levels of proteins involved in colon cancer progressions such as Survivin, EGFR, and BCL-x in colon cancer cells. Further, orally administered nanoliposomes-loaded M13 presented potent wound-healing/anti-inflammatory effects in the DSS-induced mouse model of colitis and anti-cancer effects in the AOM/DSS-induced colon cancer model. We also demonstrated that nanoliposomes-loaded M13 treatment did not affect blood. In addition, we examined the anti-inflammatory activity of M13 on active ulcerative mucosal biopsies (Reprocell tissue protocol TPS-008) from a patient (42-year-old white male) undergoing therapeutic resection for ulcerative colitis. The results of the RT2 Profiler PCR Array demonstrated that M13 had very potent anti-inflammatory activity and reduced the expression levels of the pro-inflammatory cytokines involved in IBD, including TNF-α, IL-1β, IL-6, and IL-23A, in the M13-treated human ulcerative biopsies compared to the untreated controls. Our proposed targeted therapy using nanoliposomes that deliver M13 to the colon might be used to treat ulcerative colitis and colitis-associated cancer without causing any side effects.

Antibiotic resistance, phylogenetic group, and genotyping investigation in Escherichia coli strains of gut flora in patients with colorectal cancer in Iranian population

Background: According to the World Health Organization, the prevalence of colorectal cancer (CRC) is the third-most common cancer in Iran. The population of Escherichia coli strains of the B2 phylogenetic group as one of the components of gut microbiota is increased in the gut of people with CRC and can also play a role in the process of cancerous intestinal cells or malignancy. The most common concern about treating infections and their association with cancer is multidrug resistance (MDR). Methods: In this study, stool samples of 20 patients with CRC were examined for sensitivity to 9 antibiotics and their genotypes were determined. Results: More than 90% of the isolates were MDR. Although among 20 isolates of E. coli gut flora in CRC patients, 11 (55%), 7 (35%), 0 (0%), and 2 (10%) strains belonged to phylogenetic groups B2, D, B1, and A, respectively. Conclusions: There was no significant relationship between the B2 phylogenetic group and MDR. Furthermore, the closeness of the species based on >90% similarity, seven clusters were observed that among five clusters, similarities were observed in terms of the phylogenetic group.

Potential Cytotoxic Effect of Crude Extract Produced by Lactobacillus spp on Caco-2 &amp; L20B Cell Lines in vitro

This study was designed to determine the cytotoxic effect of crude products which was produced by isolates of Lactobacillusspp against three cell lines using 3-(dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT assay). About 72samples were collected from local dairy products and infant feces with age of (1- 40 days) in Baghdad, the collection of thosesamples last long (September 2016 to January 2017). All isolates were identified as Lactobacillus spp according to their morphological,cultural, biochemical features and PCR technique by 16SrRNA gene. The cytotoxicity of crude extract was evaluatedon cancer intestinal cells of mice L20B, human colon adenocarcinoma Caco-2 and human hepatic WRL-68 cell lines. Theresults showed that crude extract had cytotoxic effect on L20B with IC50= 17 g/ml and on Caco-2 with IC50=221.9 g/ml in concentration 400 g/ml for both. While the crude extract of WRL-68 cells did not show when treated with the sameconcentration when used WRL-68 as normal cells for comparison. Conclusion: In last years, the high prevalence rate of manytypes of cancer in Baghdad, especially of intestinal cancer in human, so had used materials was produced by Probiotic bacteriato study their effects on the cancer cells in vitro.Key words: MTT assay, Caco-2 cell line, L20B cell line, Crude extracts, Lactobacillus spp.

Anti-inflammatory and anticancer effects of flavonol glycosides from Diplotaxis harra through GSK3β regulation in intestinal cells

Context and objective: Diplotaxis harra (Forssk.) Boiss. (Brassicaceae) is traditionally used as an antidiabetic, anti-inflammatory or anticancer agent. In these pathologies, the glycogen synthase kinase 3 β (GSK3β) is overactivated and represents an interesting therapeutic target. Several flavonoids can inhibit GSK3β and the purpose of this study was to search for the compounds in Diplotaxis harra which are able to modulate GSK3β.Materials and methods: Methanol extracts from D. harra flowers were prepared and the bio-guided fractionation of their active compounds was performed using inflammatory [protease-activated receptor 2 (PAR2)-stimulated IEC6 cells] and cancer (human Caco-2 cell line) intestinal cells. 50–100 μg/mL of fractions or compounds purified by HPLC were incubated with cells whose inhibited form of GSK3β (Pser9 GSK3β) and survival were analyzed by Western blot at 1 h and colorimetric assay at 24 h, respectively. LC-UV-MS profiles and MS-MS spectra were used for the characterization of extracts and flavonoids-enriched fractions, and the identification of pure flavonoids was achieved by MS and NMR analysis.Results: The methanol extract from D. harra flowers and its flavonoid-enriched fraction inhibit GSK3β in PAR2-stimulated IEC6 cells. GSK3β inhibition by the flavonoid-enriched D. harra fraction was dependent on PKC activation. The flavonoid-enriched D. harra fraction and its purified compound isorhamnetin-3,7-di-O-glucoside induced a 20% decrease of PAR2-stimulated IEC6 and Caco-2 cell survival. Importantly, normal cells (non-stimulated IEC6 cells) were spared by these treatments.Conclusion: This work indicates that flavonoids from D. harra display cytotoxic activity against inflammatory and cancer intestinal cells which could depend on GSK3β inhibition.

Annexin II binds progastrin and gastrin-like peptides, and mediates growth factor effects of autocrine and exogenous gastrins on colon cancer and intestinal epithelial cells

We and others have reported the presence of novel progastrin (PG)/gastrin receptors on normal and cancerous intestinal cells. We had earlier reported the presence of 33-36 kDa gastrin-binding proteins on cellular membranes of colon cancer cells. The goal of the current study was to identify the protein(s) in the 33-36 kDa band, and analyse its functional significance. A carbodiimide crosslinker was used for crosslinking radio-labeled gastrins to membrane proteins from gastrin/PG responsive cell lines. Native membrane proteins, crosslinked to the ligand, were solubulized and enriched by >1000-fold, and analysed by surface-enhanced laser desorption/ionization-time of flight-mass spectrometry. The peptide masses were researched against the NCBInr database using the ProFound search engine. Annexin II (ANX II) was identified, and confirmed by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry. As HCT-116 cells express autocrine PG, the in situ association of PG with ANX II was demonstrated in pulldown assays. Direct binding of PG with ANX II was confirmed in an in vitro binding assay. In order to confirm a functional importance of these observations, sense and anti-sense (AS) ANX II RNA-expressing clones of intestinal epithelial (IEC-18) and human colon cancer (HCT-116) cell lines were generated. AS clones demonstrated a significant loss in the growth response to exogenous (IEC-18) and autocrine (HCT-116) PG. We have thus discovered that membrane-associated ANX II binds PG/gastrins, and partially mediates growth factor effects of the peptides.

Expression of unprocessed, full-length progastrin, and its biological efficacy on normal and cancerous intestinal cells

Expression of unprocessed, full-length progastrin, and its biological efficacy on normal and cancerous intestinal cells

Processing and characterization of the low density lipoprotein receptor in the human colonic carcinoma cell subclone HT29-18: a potential pathway for delivering therapeutic drugs and genes.

Low density lipoprotein (LDL) processing has been investigated in the subcloned human colonic carcinoma cell line HT29-18. LDL binding at 4 degrees C was a saturable process in relation to time and LDL concentration. The Kd for LDL binding was 11 micrograms/ml. ApoE-free HDL3 or acetylated LDL did not significantly compete with 125I-LDL binding, up to 500 micrograms/ml. 125I-LDL binding was decreased by 70% in HT29-18 cells preincubated for 24 hours in culture medium containing 100 micrograms/ml unlabelled LDL. Ligand blotting studies performed on HT29-18 homogenates using colloidal gold labelled LDL indicated the presence of one autoradiographic band corresponding to an apparent molecular weight of 130 kDa, which is consistent with the previously reported molecular weight of the LDL receptor in human fibroblasts. At 37 degrees C, 125I-LDL was actively internalized by HT29-18 cells and lysosomal degradation occurred as demonstrated by the inhibitory effect of chloroquine. LDL uptake and degradation by HT29-18 cells also resulted in a marked decrease in endogenous sterol synthesis. These data demonstrate that the HT29-18 human cancerous intestinal cells are able to specifically bind and internalize LDL, and that LDL processing results in down-regulation of sterol biosynthesis. Thus, intestinal epithelial cells possess specific LDL receptors that can be exploited to accomplish drug delivery and gene transfer via the receptor-mediated endocytosis pathway.

Glycoproteins of cultured epithelial cells from human colonic adenocarcinoma and fetal intestine

The molecular weight distribution and Concanavalin A binding properties of labeled glycopeptides and glycoproteins from cell surface membranes, the cytoplasmic fraction and culture media were studied in human fetal and neoplastic colonic cells. Cells were labeled by culturing cells in the presence of [ 14C]-or[ 3 H]-labeled glucosamine or fucose. Both fucose and glucosamine were incorporated into the membrane glycoprotein fraction of the colonic cancer cells to a much lesser extent than into fetal cells. The elution profile obtained from Bio-gel P- 150 column chromatography indicated that colonic cancer cell membranes contain fucose- and glucosamine-labeled, trypsin labile, high molecular weight glycopeptides not present in fetal cells. Affinity chromatographic studies of membrane glycopeptides using agarose-bound Concanavalin A also indicated that cell surface membranes from fetal and cancerous colonic cells contain not only glycoproteins of different molecular weights, but glycoproteins having different oligosaccharide side chains. These results indicate that there are significant quantitative and qualitative differences in cell surface glycoproteins between fetal and cancerous intestinal cells.