7 research outputs found
HIC1 Expression Distinguishes Intestinal Carcinomas Sensitive to Chemotherapy
No full textAbstractNeoplastic growth is frequently associated with genomic DNA methylation that causes transcriptional silencing of tumor suppressor genes. We used a collection of colorectal polyps and carcinomas in combination with bioinformatics analysis of large datasets to study the expression and methylation of Hypermethylated in cancer 1 (HIC1), a tumor suppressor gene inactivated in many neoplasms. In premalignant stages, HIC1 expression was decreased, and the decrease was linked to methylation of a specific region in the HIC1 locus. However, in carcinomas, the HIC1 expression was variable and, in some specimens, comparable to healthy tissue. Importantly, high HIC1 production distinguished a specific type of chemotherapy-responsive tumors
Wnt Effector TCF4 Is Dispensable for Wnt Signaling in Human Cancer Cells
No full textT-cell factor 4 (TCF4), together with β-catenin coactivator, functions as the major transcriptional mediator of the canonical wingless/integrated (Wnt) signaling pathway in the intestinal epithelium. The pathway activity is essential for both intestinal homeostasis and tumorigenesis. To date, several mouse models and cellular systems have been used to analyze TCF4 function. However, some findings were conflicting, especially those that were related to the defects observed in the mouse gastrointestinal tract after Tcf4 gene deletion, or to a potential tumor suppressive role of the gene in intestinal cancer cells or tumors. Here, we present the results obtained using a newly generated conditional Tcf4 allele that allows inactivation of all potential Tcf4 isoforms in the mouse tissue or small intestinal and colon organoids. We also employed the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system to disrupt the TCF4 gene in human cells. We showed that in adult mice, epithelial expression of Tcf4 is indispensable for cell proliferation and tumor initiation. However, in human cells, the TCF4 role is redundant with the related T-cell factor 1 (TCF1) and lymphoid enhancer-binding factor 1 (LEF1) transcription factors
Unique Gene Expression Signatures in the Intestinal Mucosa and Organoids Derived from Germ-Free and Monoassociated Mice
No full textCommensal microbiota contribute to gut homeostasis by inducing transcription of mucosal genes. Analysis of the impact of various microbiota on intestinal tissue provides an important insight into the function of this organ. We used cDNA microarrays to determine the gene expression signature of mucosa isolated from the small intestine and colon of germ-free (GF) mice and animals monoassociated with two E. coli strains. The results were compared to the expression data obtained in conventionally reared (CR) mice. In addition, we analyzed gene expression in colon organoids derived from CR, GF, and monoassociated animals. The analysis revealed that the complete absence of intestinal microbiota mainly affected the mucosal immune system, which was not restored upon monoassociation. The most important expression changes observed in the colon mucosa indicated alterations in adipose tissue and lipid metabolism. In the comparison of differentially expressed genes in the mucosa or organoids obtained from GF and CR mice, only six genes were common for both types of samples. The results show that the increased expression of the angiopoietin-like 4 (Angptl4) gene encoding a secreted regulator of lipid metabolism indicates the GF status
Supplementary Materials and Methods, Figures 1 - 9, Tables 1 - 5 from Monensin Inhibits Canonical Wnt Signaling in Human Colorectal Cancer Cells and Suppresses Tumor Growth in Multiple Intestinal Neoplasia Mice
No full textPDF file - 515KB, Supplementary Materials and Methods. Supplementary Figure S1. Densitometric analysis of western blots shown in Figs. 2D (A), 3C (B) and 4D (C). Supplementary Figure S2. Monensin suppresses Wnt signaling activated by TALEN-induced disruption of the APC gene. Supplementary Figure S3. Monensin does not interfere with Dvl phoshorylation. Supplementary Figure S4. The inhibitory effect of monensin on Wnt signaling is not restricted to the vesicular acidity blockade. Supplementary Figure S5. Monensin inhibits aberrant Wnt signaling in CRC cells COLO320 and LS174T. Supplementary Figure S6. ROS concentration is not increased by monensin. Supplementary Figure S7. Membranous localization of beta-catenin in HCT116 cells. Supplementary Figure S8. Monensin does not inhibit beta-catenin acetylation. Supplementary Figure S9. Monensin does not change the fraction of proliferating cells in intestinal tumors. Supplementary Table S1. Sequences of primers used for qRT-PCR analyses. Supplementary Table S2. Monensin does not inhibit enzymatic activity of any of the tested protein kinases. Supplementary Table S3. qRT-PCR analysis of the TCF/beta-catenin target gene expression in CRC cells. Supplementary Table S4. Cell cycle analysis of SW480 and HCT116 cells. Supplementary Table S5. Analysis of phosphorylated amino acid residues in beta-catenin.</p
Supplementary Figures S1-3, Tables S1-3 from HIC1 Tumor Suppressor Loss Potentiates TLR2/NF-κB Signaling and Promotes Tissue Damage–Associated Tumorigenesis
No full textSupplementary Figures S1-3, Tables S1-3. Supplementary Figure S1. Heatmaps depicting gene expression in Hic1flox/flox MEFs treated with 4- OHT when compared to MEFs treated with vehicle (ethanol) only. Genes (299 in total) displaying significantly (q 1) in at least two time points is highlighted. Supplementary Figure S2. Loss of Hic1 results in increased counts of goblet and enteroendocrine cells. A,B, goblet (A) and enteroendocrine (B) cell distribution in the indicated segments of the small intestine. Specimens obtained from four Hic1flox/flox and four Hic1flox/flox Villin-Cre+ mice were stained using Periodic acid Schiff (PAS) and an anti-chromogranin A antibody to visualize goblet and enteroendocrine cells, respectively. Stained cells (indicated by black arrowheads in the histology images on the right) were in several different fields indicated by numbers on the X axis. C, no changes in enterocytes were noted in the Hic1-deficient small intestine. Left, qRT-PCR analysis of enterocytespecific markers hairy and enhancer of split-1 (Hes1) and sucrase-isomaltase (SI). The expression level of the respective gene in wt mice (upon normalization to Ubb) was arbitrarily set to 1. Right, staining of brush border enzyme alkaline phosphatase (AP) produced by differentiated enterocytes in the small intestine. Scale bar: 0.15 mm; error bars: SDs. Supplementary Figure S3. DSS-induced transcriptional response in the colon of Hic1flox/flox and Hic1flox/flox Villin-Cre+ mice six and nine days after DSS withdrawal. Epithelial lining of the colon obtained from four animals of each genotype was analyzed using qRT-PCR. The results were normalized to the Ubiquitin B (Ubb) housekeeping gene; the relative expression of another housekeeping gene, β-actin, is also shown. The expression level of the corresponding gene in mice without DSS treatment was arbitrarily set to 1. Error bars: SDs. Supplementary Table S1 Primers used for qRT-PCR analysis Supplementary Table S2. List of genes differentially expressed in Hic1flox/flox MEFs treated with 4- OHT for 48, 72, and 120 hours when compared to control MEFs treated (for the given time periods) with vehicle. Selection criterion: q < 0.05. Of note, none of the genes passed the selection criterion for the 24-hour time point. Supplementary Table S3. The most different 'Gene Ontology Biological Processes' (GO BPs) and WikiPathways categories in expression profiles of Hic1flox/flox MEFs 72 hours after addition of 4-OHT when compared to MEFs treated with vehicle only. In total, 268 gene probes passed the significance criterion (q-value < 0.05). Corresponding annotated genes (listed in Supplementary Table S2) were analyzed using the GO BP and WikiPathways 2015 Enricher datasets. The results were sorted according to the p-value; GO BPs and WikiPathways with p < 0.05 are listed.</p
