| [Background and Aim] MicroRNAs are a class of non-coding RNAs that function as key regulators of gene expression at the post-transcriptional level. In our previous research, we found that miR-23a was significantly up-regulated in human gastric adenocarcinoma cells. In our previous work we also choose two possible target genes for miR-23a. IRF1is a member of the interferon regulatory factor family. They have constitutive expression in many cells. IRF1plays roles in regulating a series of genes expression, and is involved in regulating cell growth and apoptosis. PPP2R5E is protein phosphatase2A, one of B56family members in the subunit B family. Protein phosphatase2A is one of the four major serine/threonine phosphatase, which is involved in negatively regulating cell growth and differentiation. IRF1and PPP2R5E are associated with cancer and have tumor suppressor activity. Expression level of IRF1and PPP2R5E is related to the malignant degree of cancer cells. IRF1and PPP2R5E decreases malignancy of gastric adenocarcinoma cells by suppressing cell growth and promoting paclitaxel-induced apoptosis. Until now, the regulation mechanism of IRF1and PPP2R5E expression is widely unknown, and microRNA (miRNA) may contribute to this process. MiRNA is an endogenetic, with18-25nucleotides in length, small non-coding RNA family. MiRNA negatively regulates expression of target gene at post-transcriptional level by binding to the3’untranslated region of target mRNA. In this study, we aim to reveal the taget gene of miR-23a in gastric adenocarcinoma, and to illustrate the regulation mechanism and the effects of the miR-23a on cellular malignancy.[Methods] Firstly, we changed the expression level of miR-23a to detect the effects on gastric adenocarcinoma cells through MTT assay, colony formation assay and TUNEL assay. And then we detected the mRNA and protein level of IRF1and PPP2R5E when miR-23a was overexpressed or inhibited in MGC803and BGC823. The miR-23a expression level and IRF1and PPP2R5E mRNA level in gastric adenocarcinoma tissues were detected by real-time quantitative PCR. In gastric adenocarcinoma cell lines MGC803and BGC823we used real-time quantitative PCR and western blot analysis to validate the over-express or knockdown vector of IRF1 and PPP2R5E. In the two cell lines we used MTT assay, colony formation test and TUNEL assay to detect the effects on cell viability, colony formation ability and paclitaxel-induced apoptosis when over-express or knock down IRFl and PPP2R5E. Then, the IRF1and PPP2R5E expression was overexpressed or inhibited accompanied by miR-23a over-expression to detect whether the changes of the malignant phenotypes caused by miR-23a could be alleviated or eliminated.[Results] miR-23a promoted cell proliferation and suppressed paclitaxel-induced apoptosis in gastric adenocarcinoma cell lines. In MGC803and BGC823cells miR-23a negatively regulated IRF1and PPP2R5E in mRNA and protein level. As shown in the result of real-time PCR, miR-23a is up-regulated whereas IRF1and PPP2R5E is up-regulated compared to the adjacent normal tissues. IRF1and PPP2R5E level is negatively associated with miR-23a in the tissue samples. In gastric adenocarcinoma tissues, IRF1and PPP2R5E are down-regulated compared to the adjacent normal tissues through immunohistochemical analysis. Our vectors are effective validated by real-time PCR and western blot, which could be used in further cell phynotype experiments. IRF1and PPP2R5E can repress cell viability and colony formation MGC803and BGC823cells, whereas promote paclitaxel-induced apoptosis of those. The alteration of these malignant phenotypes caused by overexpression of miR-23a could be alleviated or eliminated by over-expression of IRF1and PPP2R5E, respectively.[Conclusion] In gastric adenocarcinoma cells, miR-23a targetes and negatively regulates IRF1and PPP2R5E expression. miR-23a as an oncogene represses cellular apoptosis and promotes cell proliferation in gastric adenocarcinoma cells. |
PDF Full Text Request