| Gastric carcinoma is a common malignant tumor of digestive tract,The morbidity of which is particularly high in China,Japan and South Korea.Because most of patients have been diagnosed with advanced gastric cancer,chemotherapy is one of the main treatment. Chemotherapy combined with platinum as the first-line treatment for gastric cancer In clinical practice,with which has high efficacy. The resistance to platinum has become a major limitation of the therapy for the gastric cancer.However, there are a lot of studies of the resistance to platinum in recent years,but there is still lacking a method that can be use of evaluating the chemosensitivity to gastric cancer before chemotherapy.The ribonucleotide reductase (RNR) is a time-limited enzyme, which plays an important role in DNA synthesis and repair, ribonucleoside reductase 1 (RRM1) is one of the subunit of ribonucleotide reductase.higher expression of RRM1 may promote the proliferation and metastasis of tumor.the higher expression,for the patients of lung cancer in the progression with poor prognosis. however,the patients with lung adenocarcinoma of early stag could be treated surgically,prompted to a longer survival. So far,the research weather a relativity between the expression level of RRM1 in gastric cancer and the sensibility of platinum chemotherapy is still relatively less.We used the technology of gene silencing to down regulation the expression of RRM1 gene in gastric cancer of the AGS cell line,and analysed the sensitive to oxaliplatin weather Rooted in lower expression of RRM1 in this study.Objective:We try to study whether low expression of RRM1 lead to a better drug sensibility of gastric cancer cells to oxaliplatin,and to learn the relation between the RRM1 expression levels and the sensibility of gastric cancer cells to oxaliplatin.1 Methods:1.1 Cultured AGS cell line of gastric cancer.1.2 Aartificially constructed three specific interference fragments as RRM1 siRNA1,RRMl siRNA2, and RRM1 siRNA3.1.3 Used the technique of liposome-mediated transfection transfected the three interference fragments (RRM1 siRNA1,RRM1 siRNA2, RRM1siRNA3) into three groups of AGS cells respectively,as the experimental group, blank interference fragment transfection group as the negative control group and non-transfected group (bottom cell) as the blank control group.1.4 Used RT-PCR technology to detect the experimental group, negative control group and blank control group of RRM1 genes mRNA expression levels for AGS gastric cancer,and pick out the group which is best effect of RRM1 specifically interfere fragments from the three groups of experimental groups as the following experiments.1.5 Used western blotting technique to detect three experimental groups,negative control group and blank control group RRM1 relative amount of protein expression.1.6 With different ultimate concentrations of oxaliplatin for 0.2μmol/L,0.8μmol/L, 1.6μmol/L,3.2μmol/L,20μmol/L,100μmol/L treated with the experimental group, negative control group and blank control group AGS cells for 24 hours and then used the assay of CCK-8 to get the rusult and calculated the IC50 of AGS cells for each group.1.7 With ultimate concentrations of oxaliplatin for 3.0μmol/L treated with the experimental group,negative control group and blank control group AGS cells for 24 hours,and then used the flow cytometry to assay the apoptosis of the cells and calculated the apoptosis rate in each group.1.8 Used the software of SPSS 17.0 processing the data,with the statistical methods to analysis whether the lower expression of RRM1 is relating about the sensitive of gastric cancer cells to oxaliplatin.2 Results:2.1 Compared interference fragment transfected group with blank group (negative control group) and non-transfected group (control group) in gastric cancer AGS cells,the RRM1 mRNA expression levels and the relative quantity protein of the experimen group were significantly lower (P<0.05),and the interference effect of the interference fragment RRM1 siRNA3 group was the best, and which was used in the following experiments.2.2 With the treatment for different concentrations of oxaliplatin in the experimental group, negative control group and blank control group AGS cells with 0.2μmol/ L,0.8μmol/L,1.6μmol/L,3.2μmol/L,20μmol/L,100μmol/L for 24 hours,according to the CCK-8 detected experimental results,The IC50 to oxaliplatin was calculated for experimental group and blank control group and negative control of AGS cells were: 3.34μmol/L,5.85μmol/L,5.13 μmol/L (p<0.05).2.3 With the treatment of 3μmol/L oxaliplatin for AGS cells 24 hours in the experimental group,negative control group and blank control group,the apoptosis rate were (35.84 ± 2.0)%,(27.32 ± 2.6)%,(28.26 ± 1.5)%(p<0.05),respectively.3 Conclusions:3.1 transfected the RRM1 siRNA interference fragmen to the gastric AGS cells can decline expression levels of mRNA and reduce relative amount of protein expression;3.2 higher expression of RRM1 in gastric cancer is one causation of resistance to oxaliplatin,lower expression can increase the sensitivity of gastric cancer cells to oxaliplatin,so it provided a theory to reduce the amount of oxaliplatin in clinical. |
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