The Primary Study Of MiR-210 Influence On Epithelial-mesenchymal Transition

Objective: The first part of this experiment showed the effect of hypoxia on the RNA expression of HIF-1??micro RNA-210(miR-210)and EMT markers(E-cadherin,Twist1)in gastric cancer cell line MGC803.To discuss the effect of miR-210 on the epithelial mesenchymal transition(EMT)in gastric cancer cell line.The second part explored the differential expression of mi R-210 in peripheral blood of tumor patients and healthy controls and analysed the clinincal characteristics through the clinical trials.Methods:1.Choosing 200?mol/L CoCl2 to culture gastric cancer cell line MGC803 for 4h as hypoxia group(the concentration and temperature referenced to the prep work).Detecting the expression of HIF-1? miR-210 and EMT markers(E-cadherin,Twist1)through qRT-PCR,and then compared with normoxia group.2.Transient transfected mi R-210 mimics to the gastric cancer cell line MGC803 by transfection reagent Lipo 2000.Continue culturing for 24 hours after transfection.Detecting and comparing the expression of EMT markers(E-cadherin,Twist1)through qRT-PCR with control group.3.Collecting the peripheral blood of tumor patients and healthy controls.Sucking the thin-film which is rich of lymph and mononuclear cells after centrifugal.Detecting the expression of EMT markers(E-cadherin,Twist1)through the method of qRT-PCR.Comparing the difference between gastric cancer group and healthy control group.Then analyzing the relationship between the expression of miR-210 and clinincal characteristics of patients.Results:1.Hypoxia inducing gastric cancer cells for 4 hours by cobalt chloride with the concentration of 200?mol/L,efining as hypoxia group.Comparing with normoxia group,the relative expression levels of HIF-1? were 2.29±0.31vs1±0.09,the difference was statistically significant(P=0.001).The relative expression levels of miR-210 in hypoxia group and normoxia group were 3.36±0.05vs1±0.15.The relativeexpression levels of E-cadherin in hypoxia group and normoxia group were0.31±0.05vs1±0.05.The relative expression levels of Twist1 in hypoxia group and normoxia group were 1.75±0.30vs1±0.03.The difference in the each groups ware all statistically significant(P<0.05).2.The relative expression levels of miR-210 in miR-210 mimics transfection group(experience group)and control group were 678.14±54.30vs1±0.02.The relative expression levels of E-cadherin in experience group and control group were0.60±0.02vs1±0.07.The relative expression levels of Twist1 in experience group and control group were 1.24±0.04vs1±0.06.The difference in the each groups ware all statistically significant(P<0.05).3.The average levels of miR-210 expression in the peripheral blood of 23 tumor patients and 15 healthy controls were 2.21±1.60vs1±0.11,the difference was statistically significant(P=0.002).To analyze the association of the clinincal characteristics and the levels of miR-210 expression in the peripheral blood of 23 tumor patients.The results show that high expression of miR-210 is tightly related with metastasis(P=0.008).The relative expression levels of miR-210 have nothing to do with gender,age,tumor size,degree of differentiation.Conclusion:1.Hypoxia promoting the expressing of miR-210 in gastric cancer cells.Up-regulation of miR-210 may accelerate the process of EMT.2.MiR-210 may be the intermediate link of the process of hypoxia inducing EMT.3.The levels of miR-210 expression in the peripheral blood of tumor patients were significantly increased compared with healthy controls.The levels of miR-210 expression in the metastasic group were significantly increased.Showing that miR-210 may be associated with poor prognosis.