Slug and Snail have differential effects in directing colonic epithelial wound healing and partially mediate the restitutive effects of butyrate.

Restitution of wounds in colonic epithelium is essential in the maintenance of health. Microbial products, such as the short-chain fatty acid butyrate, can have positive effects on wound healing. We used an in vitro model of T84 colonic epithelial cells to determine if the Snail genes Slug (SNAI2) and Snail (SNAI1), implemented in keratinocyte monolayer healing, are involved in butyrate-enhanced colonic epithelial wound healing. Using shRNA-mediated Slug/Snail knockdown, we found that knockdown of Slug (Slug-KD), but not Snail (Snail-KD), impairs wound healing in scratch assays with and without butyrate. Slug and Snail had differential effects on T84 monolayer barrier integrity, measured by transepithelial resistance, as Snail-KD impaired the barrier (with or without butyrate), whereas Slug-KD enhanced the barrier, again with or without butyrate. Targeted transcriptional analysis demonstrated differential expression of several tight junction genes, as well as focal adhesion genes. This included altered regulation of Annexin A2 and ITGB1 in Slug-KD, which was reflected in confocal microscopy, showing increased accumulation of B1-integrin protein in Slug-KD cells, which was previously shown to impair wound healing. Transcriptional analysis also indicated altered expression of genes associated with epithelial terminal differentiation, such that Slug-KD cells skewed toward overexpression of secretory cell pathway-associated genes. This included trefoil factors TFF1 and TFF3, which were expressed at lower than control levels in Snail-KD cells. Since TFFs can enhance the barrier in epithelial cells, this points to a potential mechanism of differential modulation by Snail genes. Although Snail genes are crucial in epithelial wound restitution, butyrate responses are mediated by other pathways as well.NEW & NOTEWORTHY Although butyrate can promote colonic mucosal healing, not all of its downstream pathways are understood. We show that the Snail genes Snail and Slug are mediators of butyrate responses. Furthermore, these genes, and Slug in particular, are necessary for efficient restitution of wounds and barriers in T84 epithelial cells even in the absence of butyrate. These effects are achieved in part through effects on regulation of ß1 integrin and cellular differentiation state.

A functional polymorphism in TFF1 promoter is associated with the risk and prognosis of gastric cancer.

Trefoil Factor 1 (TFF1, also named pS2), which serves as the gastrointestinal mucosal protector, is known as gastric-specific tumor suppressor gene. However, the genetic variants of TFF1 are still not well studied. In our study, we aim to explore the effects of tagging single nucleotide polymorphisms (tagSNPs) of TFF1 on risk and prognosis of gastric cancer. Seven tagSNPs of TFF1 gene were first analyzed in the discovery set, which was consisted of 753 cases and 950 cancer-free controls. Then, the validation set (940 cases and 1,042 controls) was used for further evaluation. Moreover, we also tested the relation between these tagSNPs and prognosis of gastric cancer (GC). A series of experiments were performed to investigate the underlying mechanisms. We found that rs3761376 AA in the promoter region of TFF1, could reduce the expression of TFF1 by affecting the binding affinity of estrogen receptor 1 (ESR1, ERα), and thereby increased the risk of GC (1.29, 1.08-1.53). Moreover, the rs3761376 AA genotype was also found associated with worse prognosis among patients receiving 5-FU based chemotherapy after surgery (1.71, 1.18-2.48). Further functional assays demonstrated that TFF1 could increase the chemosensitivity of 5-FU by modulating NF-κB targeted genes. These results identified the effect of rs3761376 on TFF1 expression, which accounted for the correlation with susceptibility and prognosis of GC; and this genetic variant may be a potential biomarker to predict the risk and survival of GC.

TREFOIL FACTOR FAMILY 1 EXPRESSION CORRELATES WITH CLINICAL OUTCOME IN PATIENTS WITH RETINOBLASTOMA.

PURPOSE: Correlation of trefoil factor family 1 (TFF1) expression in retinoblastoma tumors with different clinical parameters to evaluate a potential involvement of TFF1 in tumor development and progression. METHODS: A representative cohort of 59 enucleated eyes from individual patients with retinoblastoma was analyzed for its TFF1 expression profile by immuno staining and real-time PCR. Trefoil factor family 1 expression was correlated with demographics, laterality, tumor-node-metastasis stage, International Classification of Retinoblastoma, tumor differentiation level, and treatment. RESULTS: According to our analysis, increased TFF1 expression significantly correlates with unilateral tumors diagnosed in older children and with poorly differentiated tumors and higher tumor-node-metastasis stages. CONCLUSION: This retrospective study reveals that unilateral tumors at a higher clinical tumor-node-metastasis stage and poorly differentiated tumor cells express significantly higher levels of TFF1 than those of differentiated tumors at lower tumor-node-metastasis stages. Besides, TFF1 expression correlates with the age of the patients at the time of tumor diagnosis. Our data indicate that TFF1 expression levels are potentially useful additional markers in the classification of tumor staging and prognosis of patients with retinoblastoma.

Differences between gastric signet-ring cell carcinoma and poorly differentiated adenocarcinoma: A comparison of histopathologic features determined by mucin core protein and trefoil factor family peptide immunohistochemistry.

We investigated differences between the pathological features of gastric signet-ring cell carcinoma (sig) and poorly differentiated adenocarcinoma (por) by examining the expressions of the trefoil factor family peptides (TFFs) and mucin core proteins (MUCs). Ninety-seven tissues of 97 gastric cancer patients were selected for this study. After gastrectomy, the major histopathologic types were determined to be sig, solid-type poorly differentiated adenocarcinoma (por1), non-solid type poorly differentiated adenocarcinoma (por2), and well-differentiated tubular adenocarcinoma (tub1). We evaluated the prevalence of positive staining for MUCs (MUC5AC and MUC2) and TFFs (TFF1 and TFF3) and assessed the correlation between MUCs and TFFs in each histopathological type. The rate of MUC2 expression significantly differed between sig and por2 (50.0% vs 11.7%, P = 0.011). TFF3 expression in sig significantly differed from TFF3 expression in both por2 (100% vs 17.6%, P < 0.0001) and por1 (100% vs 33.3%, P = 0.0004). MUC5AC and TFF1 expressions were significantly correlated in por1 (r = 0.705, P = 0.002), por2 (r = 0.535, P = 0.0009), and tub1 (r = 0.470, P = 0.0034), while MUC2 and TFF3 expressions were significantly correlated only in sig (r = 0.593, P = 0.040). The expression and correlation patterns of the TFFs and MUCs suggest that the histopathologic features of gastric sig differ from those of por.

Inducible gene modification in the gastric epithelium of Tff1-CreERT2, Tff2-rtTA, Tff3-luc mice.

Temporal and spatial regulation of genes mediated by tissue-specific promoters and conditional gene expression systems provide a powerful tool to study gene function in health, disease, and during development. Although transgenic mice expressing the Cre recombinase in the gastric epithelium have been reported, there is a lack of models that allow inducible and reversible gene modification in the stomach. Here, we exploited the gastrointestinal epithelium-specific expression pattern of the three trefoil factor (Tff) genes and bacterial artificial chromosome transgenesis to generate a novel mouse strain that expresses the CreERT2 recombinase and the reverse tetracycline transactivator (rtTA). The Tg(Tff1-CreERT2;Tff2-rtTA;Tff3-Luc) strain confers tamoxifen-inducible irreversible somatic recombination and allows simultaneous doxycycline-dependent reversible gene activation in the gastric epithelium of developing and adult mice. This strain also confers luciferase activity to the intestinal epithelium to enable in vivo bioluminescence imaging. Using fluorescent reporters as conditional alleles, we show Tff1-CreERT2 and Tff2-rtTA transgene activity in a partially overlapping subset of long-term regenerating gastric stem/progenitor cells. Therefore, the Tg(Tff1-CreERT2;Tff2-rtTA;Tff3-Luc) strain can confer intermittent transgene expression to gastric epithelial cells that have undergone previous gene modification, and may be suitable to genetically model therapeutic intervention during development, tumorigenesis, and other genetically tractable diseases. Birth Defects Research (Part A) 106:626-635, 2016. © 2016 Wiley Periodicals, Inc.