9966

TNFSF15

Intestinal subepithelial myofibroblasts (SEMFs) exert a profibrotic role in Crohn's disease (CD). tumor necrosis factor-like cytokine 1A (TL1A) and its receptors, death-domain receptor 3 (DR3) and decoy receptor 3 (DcR3), are mucosal factors with significant involvement in experimental inflammation and CD. We aimed to determine the regulation of expression of this system of proteins in SEMFs and intestinal epithelial cells. The relative amount of mRNA transcripts for TL1A, DR3, and DcR3 was measured by real-time reverse transcription polymerase chain reaction in cultured primary SEMFs, colonic myofibroblast cell line 18CO, and epithelial cell line HT29. Protein expression was determined by immunofluorescence. The effect of various proinflammatory stimuli in mRNA and protein expression was studied. TL1A mRNA and protein expression in primary SEMFs (and 18CO cells) was significantly upregulated after stimulation with interleukin 1-alpha and/or tumor necrosis factor alpha (TNF-alpha) (32- to 44-fold increase, P < 0.05 vs unstimulated). Following stimulation with interleukin 1-alpha + TNF-alpha + IFN-gamma, HT-29 cells highly expressed DR3 (4.1-fold over unstimulated, P = 0.008) and DcR3 (56-fold, P = 0.009) and secreted soluble factors that led to induction of TL1A mRNA in primary SEMFs (28-fold, P = 0.008). Activated epithelial cells significantly upregulated IL-8 expression in response to stimulation with recombinant TL1A. Supernatants from mucosal cultures of patients with CD were able to stimulate the expression of TL1A in cultured primary SEMFs, in comparison to supernatants from healthy controls (3.8-fold increase, P < 0.05) or culture media alone (P < 0.05). In conclusion, we found that proinflammatory cytokines are important regulators of the expression of TL1A in SEMFs and of its receptors in intestinal epithelial cells. Our results raise the possibility for involvement of TL1A/DR3/DR3-mediated mechanisms in epithelial-mesenchymal interactions and the development of inflammation-induced intestinal fibrosis in CD.CI - Copyright (c) 2016 Elsevier Inc. All rights reserved.

The present study was carried out to investigate the effects of vascular endothelial growth inhibitor 174 (VEGI174) and its functional domains (V7 and V8) on epithelialmesenchymal transition (EMT) in renal cell carcinoma (RCC) cells in vitro. The RCC cell lines A498 and 786O were used in this study. Based on our preliminary study, we selected fulllength VEGI174 and its functional domains (V7 and V8) as the target genes in this study. Plasmids containing VEGI174, V7 or V8 transgenes were constructed and transfected into A498 and 786O cell lines. Cytological activity was tested during cell culture. Quantitative PCR and western blot analysis were performed to determine the expression levels of EMT markers (Ecadherin, vimentin, betacatenin and Slug). Overexpression of VEGI174, V7 or V8 did not have a significant influence on cell viability (P>0.05). The mRNA level of Ecadherin was significantly upregulated, while that of vimentin was downregulated in A498VEGIexp, A498V7exp, A498V8exp, 786OVEGIexp, 786OV7exp and 786OV8exp cells compared with the cells containing the empty plasmid controls (P<0.05). The western blot results showed that changes in protein expression levels were consistent with the changes in mRNA expression. Both the mRNA and protein expression levels of betacatenin and Slug were downregulated in the A498VEGIexp, A498V7exp, A498V8exp, 786OVEGIexp, 786OV7exp and 786OV8exp cells. In conclusion, overexpression of VEGI174, V7 or V8 inhibited EMT in A498 and 786O cells. Notably, V7 and V8 are two effective functional domains of VEGI174 that have the potential to be studied for peptide synthesis and the treatment of RCC.

BACKGROUND: Vascular endothelial growth inhibitor (VEGI) is a member of the tumor necrosis factor superfamily, identified as an anti-angiogenic cytokine. However, the effect of VEGI on epithelial-mesenchymal transition (EMT) in renal cell carcinoma (RCC) is still unknown. MATERIALS AND METHODS: In this study, protein VEGI174 was designed and synthesized. Renal cell carcinoma A498 cells were implanted into immune-deficient mice to establish tumor models. Two groups were included: control group treated with saline, and VEGI174-treated group. Data of tumor growth were collected every 3 to 4 days. Two weeks later, the tumor specimens were harvested for immunohistochemical staining of EMT markers (E-cadherin, N-cadherin, vimentin). RESULTS: Compared to the saline-treated group, the VEGI174-treated group showed significant inhibition of tumor growth (p<0.05). The expression of E-cadherin was significantly higher in the VEGI174-treated group compared to the saline-treated group (p<0.01). However, the expression of N-cadherin and vimentin were reduced in the VEGI174-treated group. CONCLUSION: Our findings indicate that VEGI174 prevents progression and tumor metastasis through inhibiting EMT in RCC in vivo. This may provide a new approach for the treatment of RCC.CI - Copyright(c) 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.