| Objective: To investigate the effect of mi R-200 c on gastric cancer cells by detecting the expression of miR-200 c in different tumor cell lines;to determine the relations between epithelial-mesenchymal transition and gastric cancer cell resistance and the role of mi R-200 c in both by detecting related indicators.Method: The expression level of mi R-200 c in varying degrees of differentiation of gastric cancer cells was measured by RT-qPCR technology to explore the role of mi R-200 c. And then,Objective gastric cancer cell line SGC-7901 was screened according to the expression level of mi R-200 c in varying degrees of differentiation of gastric cancer cells. For the gastric cancer cell SGC-7901 and adriamycin-resistant gastric cancer cell SGC-7901 / ADR, the cell inhibition rate of the two group gastric cancer cells was measured using a modified MTT assay and using Westernblot measure the expression of the protein of the MRP-1 to prove the gastric cancer cell SGC-7901 / ADR resistant. Then the cell morphology of the gastric cancer cell SGC-7901 / ADR was observed, the ability to invade was measured by scratch-wound assay, the RT-qPCR technology was used to assay the expression level of miR-200 c and the expression of Snail,Twist1 gene and the expression of protein E-Cadherin was determined with the Western-blot protein assay method to explore the relevance of the resistance and epithelial-mesenchymal transformation(EMT). Followed, the sequence of Has-mi R-200 c mimics and Has-miR-200 c inhibitor transfected into the gastric cancer SGC-7901 / ADR using Lipofectamine 2000 reagent.The cells were divided into two groups: the experimental group was transfected with Has-miR-200 c mimics and Has-miR-200 c inhibitor of gastric cancer cell SGC-7901 / ADR, the blank group was the gastric cancer cell SGC-7901 / ADR. And then, we used RT-qPCR to measure the transfection efficiency of the miR-200 c and the expression of Snail, Twist1 gene,Western-blot to detect the expression of protein E-Cadherin and MRP-1, MTT to assay the cell inhibition rate of the three group gastric cancer cells.Result: The expression level of miR-200 c was different in varying degrees of differentiationof gastric cancer cell lines. The higher the expression level, the more the degree of expression. For the gastric cancer cell SGC-7901 and the adriamycin-resistant gastric cell SGC-7901 / ADR, in the case of the adriamycin-resistant cell SGC-7901 / ADR had obvious resistance confirmed by MTT assay and western blot, we observed the cell morphology of the adriamycin-resistant gastric cell SGC-7901/ADR was spindle-like. Compared the gastric cancer cell SGC 7901, the gastric cancer cell SGC-7901/ADR had a larger cell spacing, arrangement and polymeric loose, shape with relatively large sample, obvious pseudopodia, which meet the morphological characteristics of EMT; compared with the gastric cancer cell SGC-7901, the gastric cancer cell SGC-7901/ADR had a stronger invasive ability under the same culture conditions and time by scratch-wound assay;the protein of the E-Cadherin decreased obviously and the EMT feature genes of Snail1 and Twist1 increased significantly, which meeted the molecular change of the EMT. With the RT-qPCR assay method, compared with the gastric cancer cell SGC-7901, the expression level of mi R-200 c of the gastric cancer cell SGC-7901/ADR was less and about 0.4 times, which had statistically significant(P <0.05). In the case of the gastric cancer cell SGC-7901/ADR transfected with Has-miR-200 c mimics and Has-miR-200 c inhibitor, compared with the Blank group,the expression level of miR-200 c respectively increased approximately 4.7-fold and 2.5-fold decreases, which was statistically significant(P <0.05). Each group inhibition rate from MTT assay, we could get that the inhibition rate of the gastric cancer cell SGC-7901/ADR transfected Has-miR-200 c mimics was highest, the blank group middle and the gastric cancer cell SGC-7901/ADR transfected Has-miR-200 c inhibitor lowest. The expression levels of the gene of Snail1 and Twist1 decreased significantly in the gastric cancer cell SGC-7901/ADR transfected Hsa-miR-200 c mimics and increased significantly in the gastric cancer cell SGC-7901/ADR transfected Has-miR-200 c inhibitor. The protein of the E-Cadherin significantly increased in the gastric cancer cell SGC-7901/ADR transfected Has-miR-200 c mimics and obviously decreased in the gastric cancer cell SGC-7901/ADR transfected Has-miR-200 c inhibitor. While, the expression of the protein of the MRP1 was converse of the protein of the E-Cadherin in the gastric cancer cell transfected Has-miR-200 c mimics and Has-miR-200 c inhibitor.Conclusion: The expression level of mi R-200 c is different in varying degrees of differentiation of gastric cancer cells. The gastric cancer cell with drug resistance has a tendency to epithelial-mesenchymal transformation. The mi R-200 c regulates the gastric cancer cell to resistance and the role of the occurrence of EMT and is negatively correlated with the drug resistance and the EMT. |
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