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The Effect of miR-214 on Invasion of Gasric Cancer Cell Line SGC7901

Author: ZhaoXueZuo
Tutor: ZhengChangLi
School: Central South University
Course: Pathology and Pathophysiology
Keywords: microRNA miR-214 Gastric cancer Attack Migrate PTEN
CLC: R735.2
Type: Master's thesis
Year: 2011
Downloads: 97
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Abstract

Background: Gastric cancer is a common malignancy, diagnosis and treatment of gastric cancer has made some progress, but the prognosis of patients with gastric cancer is still not satisfactory, invasion and metastasis is the main reason for its poor prognosis. The small RNA (microRNAs, miRNAs) is one of the most attention in the cancer research of biological macromolecules. The study found that miRNA play an important role in tumor metastasis and infiltration which, by inhibiting the expression of target genes involved in extracellular matrix degradation, epithelial mesenchymal transition (EMT) and tumor angiogenesis, invasion and metastasis of the tumor . Gastric cancer invasion and metastasis of miRNAs study is relatively small, Japanese scholars of the 353 patients with gastric cancer: miRNAs expression profiling analysis found that miR-214 is highly expressed in the depth of tumor invasion, invasion, lymph node metastasis and staging an independent risk factor. Our previous study also found that gastric cancer cell line BGC823, MKN45 and SGC7901 in miR-214 expression levels than normal gastric mucosal cell line GES-1 increased. About the research: miR-214 target genes and mechanisms found in the inflammatory response, miR-214 inhibit its transcription and translation by PTEN mRNA 3'-untranslated region combined, people mononuclear macrophage cell line Li et al. THP-1 apoptosis delay. Jindra found in T-cell activation, miR-214 expression was elevated, and a negative correlation with the level of expression of PTEN. The study also found that miR-214 in ovarian cancer can inhibit transcription of PTEN translation, activation of PI3K/AKT pathway, inhibition of tumor cell apoptosis induced by cisplatin resistance. MiR-214 is indeed the invasion and metastasis, needs a large number of experimental studies confirmed that its mechanism of action remains to be elucidated. Objective: To investigate the miR-214 on the invasion and migration ability of SGC7901 cells and its possible mechanism of action. Method: 1. Applications (?) MiRanda leather Pakistan due to the pre-brew measured website and TarBase of database analysis of PTEN is the target genes of miR-214 and to determine PTEN mRNA and miR-214 binding sites. Were transiently transfected Mimics the best choice of IniR-214 inhibitors and unrelated sequences into SGC7901 cell lines, to unrelated sequence transfected group (NC) as a negative control, blank transfection group (control group) for the blank control, transfection 24h after the transfection efficiency was observed under a fluorescence microscope. 3. Real-time PCR method to detect different the transfection group of SGC7901 cells the expression level of miR-214. 4 Western blot method to compare different transfection group SGC7901 cells PTEN protein expression. Transwell chamber Transwell migration assay and cell scratch assay transfection Mimics the best choice of miR-214 inhibitors SGC7901 cell invasion Migration Competence. Results: 1. Applications TarBase, miRanda found that PTEN is one of the target genes of miR-214 and miR-2145'-seed sequence region and PTEN mRNA 3'-UTR conserved sequences contiguous complementary base combine to form the miRNA : mRNA dimer has a certain stability. 24h after transfection were counted under a fluorescence microscope, miR-214 mimics and inhibitors transfected human gastric cancer SGC7901 cells, the transfection efficiency of more than 80% 3. 24h after transfection, Real-time PCR detection of different transfection group The relative expression level of miR-.214 Mimics the best choice group and inhibitors of miR-214 cells transfected with the expression level of miR-214 are the control group of 82,902.31, 0.16 times. Western blot results: SGC7901 cells transfected with miR-214 mimics, PTEN protein relative expression levels (PTEN / GAPDH grayscale ratio, 0.049 ± 0.007), independent of the sequence group (0.087 ± 0.007) than transfected, transfected with miR- of 214 inhibitors group (0.093 ± 0.010) and blank transfection group (0.099 ± 0.012) lower, the difference was statistically significant (P lt; 0.05). In Transwell invasion chamber experimental results: transfected miR-214 mimics group, the number of invasive cells was 43.60 ± 4.39 the unrelated sequence significantly higher than the group (25.80 ± 7.79), transfection of miR-214 inhibitors group (16.60 ± 5.32) and blank turn transfected group (24.60 ± 6.54), the difference was statistically significant (P lt; 0.001). MiR-214 inhibitors group transfected invasion cell number and sequence group, blank transfection group compared, the difference was not statistically significance (P gt; 0.05). 6. Transwell migration assay results: Transfection of miR-214 mimics group penetrating cell number for 39.00 ± 6.04, the unrelated sequence significantly higher than the group (26.20 ± 2.39), transfection of miR-214 inhibitors group (20.00 ± 2.55) and blank turn transfected group (20.40 ± 4.77), the difference was statistically significant (P lt; 0.05). Transfected miR-214 inhibitors group wearing the membrane cell number and sequence group, blank transfected group compared the difference was not statistically significance (P gt; 0.05) cell scratch assay results: different scratches healing time point, the the scratch width transfected the miR-214mimics group with three other transfected group the same point in time scratch width narrower, the difference was statistically significant (P lt; 0.05) transfection factors main factorial analysis results are shown effect (F = 34.913, P lt; 0.01), healing time main effect (F = 417.910, P lt; 0.01) and the interaction of two factors (F = 8.520, P lt; 0.01) were statistically significant. Conclusion: 1.miR-214 can increase the invasion and migration ability of SGC7901 cells. 2.miR-214 may promote gastric cancer cell invasion and migration through inhibition of PTEN expression.

The Effect of miR-214 on Invasion of Gasric Cancer Cell Line SGC7901

Abstract

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