human primary colonic epithelial cells (hcec)  (Applied Biological Materials Inc)

Journal: Cancers

Article Title: Intestinal Epithelial Cells Adapt to Chronic Inflammation through Partial Genetic Reprogramming

doi: 10.3390/cancers15030973

Figure Lengend Snippet: Human colonic epithelial cells adapt to chronic inflammation-associated oxidative stress in vitro. ( A ) HCEC-hTERT cells (left panel) were subjected to chronic inflammation by culturing them in the presence of activated macrophage supernatant (+AMS) or tert-Butyl-hydroperoxide (TBHP). Cells committed to senescence after a week, demonstrated by the detection of SA-β-galactosidase activity (central panel), and maintained their senescent phenotype for three weeks before resuming proliferation (right panel). ( B ) Assessment in both experimental settings (left AMS, right TBHP) of phospho-serine 10 histone H3 (pS10-H3), p53, acetylated-p53 (Ac-p53), and p21 CIP1 (p21) by Western blot (protein levels were quantified relative to the β-actin and HCEC-hTERT cells) and of SA-β-galactosidase-positive cells in parental HCEC-hTERT cells (1) and senescent cells (2). A non-parametric Student’s t -test, mean, and SD of one experiment performed in triplicate are shown. *** p < 0.001, ** p < 0.01. Uncropped Western blots are provided in .

Article Snippet: Human primary colonic epithelial cells (HCEC) were purchased from Applied Biological Materials (abm, Richmond, Canada) and immortalized through transduction of hTERT (HCEC-hTERT).

Techniques: In Vitro, Activity Assay, Western Blot

Journal: Cancers

Article Title: Intestinal Epithelial Cells Adapt to Chronic Inflammation through Partial Genetic Reprogramming

doi: 10.3390/cancers15030973

Figure Lengend Snippet: Escape from chronic-inflammation-induced senescence does not rely on p53 inactivation. ( A ) Confirmation of the wild-type status of the TP53 gene in HCEC-hTERT cells and their derivatives. TP53 exons within parental HCEC-hTERT cells and their Esc-Inf derivatives (of two independent experiments, Esc-Inf A and B) were fully sequenced. No mutation in TP53 was found, as shown by the sequences encompassing the exons encoding the residues R175, R248, and R273 that constitute the main mutation hot-stops reported in colitis-associated colon cancers. ( B ) The functionality of the p53 pathway in both parental HCEC-hTERT and its Esc-Inf derivatives was compared by submitting cells to an increasing dose of Fluorouracil (5-FU) over 24 h. The phosphorylation of the Ser10 of p53 (pS15-p53) and the levels of p53 and p21 CIP1 (p21) were assessed by Western blot. Protein levels were quantified relative to the β-actin and HCEC-hTERT cells. ( C ) The ectopic expression of p53 variants does not facilitate the escape of HCEC-hTERT cells from chronic-inflammation-induced senescence. Upper panels: cells were successively infected with mutant p53 retroviral-expressing vectors (as indicated on top), cultured in the presence of activated macrophage supernatants (+AMS) for ten days, and stained with crystal violet. Esc-Inf cells were used as an internal positive control. Percentages of stained surface are indicated. Lower panel: analysis of p53 by Western blot. Protein levels were quantified relative to the β-actin and HCEC-hTERT cells. Uncropped Western blots are provided in .

Article Snippet: Human primary colonic epithelial cells (HCEC) were purchased from Applied Biological Materials (abm, Richmond, Canada) and immortalized through transduction of hTERT (HCEC-hTERT).

Techniques: Mutagenesis, Phospho-proteomics, Western Blot, Expressing, Infection, Retroviral, Cell Culture, Staining, Positive Control

Journal: Cancers

Article Title: Intestinal Epithelial Cells Adapt to Chronic Inflammation through Partial Genetic Reprogramming

doi: 10.3390/cancers15030973

Figure Lengend Snippet: Escape from chronic-inflammation-induced or TBHP-induced senescence relies on the induction of ZEB genes. ( A ) Relative expression of SNAI1 , TWIST2 , ZEB1, and ZEB2 in cells that escaped from chronic-inflammation- (left panel) or TBHP-induced senescence (right panel), as assessed by qRT-PCR. Levels are expressed relative to the housekeeping HPRT1 gene and are normalized against HCEC-hTERT cells. One sample t and Wilcoxon test and the mean and SD of three independent experiments are shown. * p < 0.05 Central panel: analysis of ZEB1 in Esc-Inf cells by Western blot. EMT-committed Hs578T breast cancer cells were used as a positive protein detection control. Protein levels were quantified relative to the α-tubulin and Esc-Inf cells. ( B ) Ectopic expression of murine Zeb1 or Zeb2 facilitates escape from chronic inflammation-induced (+AMS, left panels) or chemical, peroxide-induced (+TBHP, right panels) senescence, as assessed in a crystal violet assay. Percentages of stained surface are indicated. Uncropped Western blots are provided in .

Article Snippet: Human primary colonic epithelial cells (HCEC) were purchased from Applied Biological Materials (abm, Richmond, Canada) and immortalized through transduction of hTERT (HCEC-hTERT).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control, Crystal Violet Assay, Staining

Journal: Cancers

Article Title: Intestinal Epithelial Cells Adapt to Chronic Inflammation through Partial Genetic Reprogramming

doi: 10.3390/cancers15030973

Figure Lengend Snippet: The ZEB-driven resistance relies on a partial genetic reprogramming. ( A ) Venn diagram highlighting the number of genes commonly up- or down-regulated (cut-off > 1.5) in HCEC-hTERT cells which had either escaped from chronic-inflammation-induced senescence (Esc-Inf A or B from two independent experiments), TBHP-induced senescence (Esc-TBHP A or B from two independent experiments), or ectopically expressed either Zeb1 or Zeb2 (HCEC- Zeb 1 and HCEC- Zeb 2) when compared to their HCEC-hTERT parental counterparts. Common up- and down-regulated genes constitute the basis for the OSAP program. ( B ) Listing of the genes commonly up-regulated (in red) or down-regulated (in green) in Esc-Inf (A and B), Esc-TBHP (A and B), HCEC- Zeb1 , and HCEC- Zeb2 compared to the parental HCEC-hTERT cells (cut-off ≥ 1.5). ( C ) Expression analysis of APOE and QPRT in cells that escaped either from chronic-inflammation-induced (upper panels) or TBHP-induced (lower panels) senescence. Left panels: expression analysis by qRT-PCR. Levels are expressed relative to the housekeeping HPRT1 gene and are normalized against HCEC-hTERT cells. One sample t and Wilcoxon test and the mean and SD of three independent experiments are shown. Right panels: analysis of APOE and QPRT by Western blot. Protein levels were quantified relative to the β-actin and HCEC-hTERT cells. ** p < 0.01, * p < 0.05. Uncropped Western blots are provided in .

Article Snippet: Human primary colonic epithelial cells (HCEC) were purchased from Applied Biological Materials (abm, Richmond, Canada) and immortalized through transduction of hTERT (HCEC-hTERT).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Cells

Article Title: Crocin Inhibits Angiogenesis and Metastasis in Colon Cancer via TNF-α/NF-kB/VEGF Pathways

doi: 10.3390/cells11091502

Figure Lengend Snippet: Cytotoxic effect of crocin on Human carcinoma cells. ( A ) HT-29 cells; ( B ) Caco-2 cells; ( C ) Human colonic epithelial cells (HCEC) and ( D ) Human umbilical vein endothelial cells (HUVEC). Values represent mean ± SD, n = 3. Asterisks indicate significant difference compared to control (* p < 0.05, ** p < 0.01), ns = non-significant, Doxorubicin (100 µg/mL) was used as positive control. The control was treated with 1% DMSO. (PC = positive control).

Article Snippet: Colon cancer cell lines (HT-29 and Caco-2 cells) and human colonic epithelial cells (HCEC) were purchased from American type culture collection.

Techniques: Control, Positive Control