BACKGROUND Gastric carcinogenesis can be induced by chronic inflammation triggered by Helicobacter pylori ( H. pylori ) infection. Tumor necrosis factor (TNF)-α and its receptors (TNFR1 and TNFR2) regulate important cellular processes, such as apoptosis and cell survival, and the disruption of which can lead to cancer. This signaling pathway is also modulated by microRNAs (miRNAs), altering gene expression. AIM To evaluate the mRNA and miRNAs expression involved in the TNF-α signaling pathway in gastric cancer (GC) tissues and its relationship. METHODS Quantitative polymerase chain reaction (qPCR) by TaqMan ® assay was used to quantify the RNA transcript levels of TNF-α signaling pathway ( TNF, TNFR1, TNFR2, TRADD, TRAF2, CFLIP, NFKB1 , NFKB2, CASP8, CASP3) and miRNAs that targets genes from this pathway (miR-19a, miR-34a, miR-103a, miR-130a, miR-181c) in 30 GC fresh tissue samples. Molecular diagnosis of H. pylori was performed by nested PCR for gene HSP60 . A miRNA:mRNA interaction network was construct using Cytoscape v3.1.1 from the in silico analysis performed using public databases. RESULTS Up-regulation of cellular survival genes as TNF, TNFR2, TRADD, TRAF2 , CFLIP , and NFKB2 , besides CASP8 and miR-34a was observed in GC tissues, whereas mediators of apoptosis such as TNFR1 and CASP3 were down-regulated. When the samples were stratified by histological type, the expression of miR-103a and miR-130a was significantly increased in the diffuse-type of GC compared to the intestinal-type. However, no influence of H. pylori infection was observed on the expression levels of mRNA and miRNAs analyzed. Moreover, the miRNA:mRNA interaction network showed several interrelations between the miRNAs and their target genes, highlighting miR-19a and miR-103a, which has as predicted or validated target a large number of genes in the TNF-α pathway, including TNF, TNFR1, TNFR2, CFLIP, TRADD, CASP3 and CASP8 . CONCLUSION Our findings show that cell survival genes mediated by TNF/TNFR2 binding is up-regulated in GC favoring its pro-tumoral effect, while pro-apoptotic genes as CASP3 and TNFR1 are down-regulated, indicating disbalance between apoptosis and cell proliferation processes in this neoplasm. This process can also be influenced by an intricate regulatory network of miRNA:mRNA.
Trypanosoma cruzi presents a high degree of intraspecific variability, with possible implications for the pathogenesis of Chagas disease. The aim of this study was to evaluate T. cruzi kDNA minicircle gene signatures using the low-stringency single-specific-primer PCR technique in both peripheral blood and oesophageal mucosa from chronic chagasic patients, with or without megaesophagus, alone or in combination with cardiopathy and megacolon. It was not possible to identify a uniform pattern of shared bands between blood and oesophageal mucosa samples from individuals with the same clinical form or mixed forms, suggesting multiple T. cruzi infections with differential tissue tropism. Thus, the results indicate that there is an intense intraspecific variability in the hypervariable regions of T. cruzi kDNA, which has so far made it impossible to correlate the genetic profile of this structure with the clinical manifestations of Chagas disease.
BACKGROUND Chronic inflammation due to Helicobacter pylori ( H. pylori ) infection promotes gastric carcinogenesis. Tumour necrosis factor-α (TNF-α), a key mediator of inflammation, induces cell survival or apoptosis by binding to two receptors (TNFR1 and TNFR2). TNFR1 can induce both survival and apoptosis, while TNFR2 results only in cell survival. The dysregulation of these processes may contribute to carcinogenesis. AIM To evaluate the effects of TNFR1 and TNFR2 downregulation in AGS cells treated with H. pylori extract on the TNF-α pathway. METHODS AGS cell lines containing TNFR1 and TNFR2 receptors downregulated by specific shRNAs and nonsilenced AGS cells were treated with H. pylori extract for 6 h. Subsequently, quantitative polymerase chain reaction with TaqMan ® assays was used for the relative quantification of the mRNAs ( TNFA, TNFR1, TNFR2, TRADD, TRAF2, CFLIP, NFKB1, NFKB2, CASP8, CASP3 ) and miRNAs (miR-19a, miR-34a, miR-103a, miR-130a, miR-181c) related to the TNF-α signalling pathway. Flow cytometry was employed for cell cycle analysis and apoptosis assays. RESULTS In nonsilenced AGS cells, H. pylori extract treatment increased the expression of genes involved in cell survival and inhibited both apoptosis ( NFKB1, NFKB2 and CFLIP ) and the TNFR1 receptor. TNFR1 downregulation significantly decreased the expression of the TRADD and CFLIP genes, although no change was observed in the cellular process or miRNA expression. In contrast, TNFR2 downregulation decreased the expression of the TRADD and TRAF2 genes, which are both important downstream mediators of the TNFR1-mediated pathway, as well as that of the NFKB1 and CFLIP genes, while upregulating the expression of miR-19a and miR-34a . Consequently, a reduction in the number of cells in the G0/G1 phase and an increase in the number of cells in the S phase were observed, as well as the promotion of early apoptosis. CONCLUSION Our findings mainly highlight the important role of TNFR2 in the TNF-α pathway in gastric cancer, indicating that silencing it can reduce the expression of survival and anti-apoptotic genes.
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