The Significance And Function Of MicroRNA-1228*Expression In Gastric Cancer

Background:miRNA are a class of short (20-23nucleotides in length), endogenous, single-stranded RNA which regulate gene expression by causing translational repression or mRNA degradation. Dysregulated miRNA play critical roles during carcinogenesis and cancer progression. Gastric cancer is the fourth most common malignant tumor worldwide and currently is the second leading cause of cancer-related mortality. Although current practice includes incorporating chemotherapy or radiation into surgical resection treatment protocols, gastric cancer survival rates remain poor. The purpose of this study was to elucidate the role of miR-1228*in gastric cancer and its corresponding molecular mechanisms.Methods:The expression level of the mature miR-1228*was examined using TaqMan real-time PCR in50pairs of human gastric cancer tissues and pair-matched adjacent noncancerous gastric tissues. Furthermore, miR-1228*expression were evaluated with regards to the clinicopathological characteristics of the patients. To evaluate the potential effects of miR-1228*on gastric cancer cells growth in vivo, stable transfection of SGC-7901cells with miR-1228*or miR-NC were injected subcutaneously into nude mice. The volume of each tumor was measured twice each week. The mice were sacrificed after5weeks and tumors from each group were weighed immediately after removal. In order to validate the hypothesis that NF-kB activation may be attributed to the downregulation of miR-1228*in gastric cancer, SGC-7901cells were transiently transfected with pGL3-1228*-promoter or pGL3-control, TNF-a was added to medium and incubated for8hours, then measured relative luciferase activity. Furthermore, we cotransfected c-Rel and miR-1228*promoter to the SGC-7901cells and measured relative luciferase activity after48hours. To further test the influence of miR-1228*on the "NF-κB signalling pathway, NF-κB Reporter constructs were transfected into either miR-1228*or miR-NC stable transfected SGC-7901cells, respectively. Western blot analysis of epithelial marker E-cadherin and mesenchymal marker Vimentin in miR-1228*stable transfected SGC-7901cells were measured. To confirming the inhibition of EMT, stably transfected SGC-7901cells transwell assay was used.Results:miR-1228*is frequently down-regulated in human gastric cancer tissues. And there was a significant correlation between miR-1228*expression and tumor size. Upregulation of the miR-1228*significantly decreased tumor growth as assessed by measurements of tumor volume than that of the control. The wet weight of tumors with miR-1228*overexpression was also significantly lower. TNF-a and c-Rel could inhibit miR-1228*promoter vector luciferase expression. The cellular transcriptional activity of NF-κB was significantly decreased on miR-1228*overexpression. miR-1228*downregulated Vimentin and upregulated E-cadherin of SGC-7901. Transwell migration assays showed that SGC-7901cells stable transfected with miR-1228*had lower migratory potentials in compare with miR-NC.Conclusion:miR-1228*is frequently down-regulated in human gastric cancer tissues. Overexpression of miR-1228*inhibits gastric cancer cells growth in nude mice. We identify a double negative feedback loop between miR-1228*and activation. miR-1228*acts as an EMT suppressor in gastric cancer cells.