| Research Objective:Based on the research of this subject,it was found that the expression of a variety of miRNA(Micro RNA)in Burkitt lymphoma was up-regulated or down-regulated,The role of miRNA in cell proliferation,differentiation and apoptosis and the key role of miR-155 in tumors.The expression of miR-155 in Burkitt lymphoma was detected,and the effect of inhibition and over-expression of miR-155 on cell proliferation and cell protein was detected,for further study of miRNA and miR-155 in Burkitt lymphoma Role provides research foundation.Research methods:After cell culture of Burkitt lymphoma EBV(Epstein-Barr virus)positive cells(EBV~+Raji)and EBV negative cells(EBV~-Ramous),Sybgreen fluorescence quantitative PCR was used to detect the expression of miR-155 in the cells.Using Lipofectamine 2000 liposome method to complement miR-155inhitor(miR-155-inhibitor sequence is complementary to mature miR-155 is a chemically modifiedsingle-chain molecule that selectively inhibits the function of miR-155)and its negative control sequence respectively Transfect EBV~+Raji cells,namely miR-155-inhibitor(Raji-in),Raji-yin(Raji-CON),miR-155-mimics(miR-155-mimics sequebce is a double-stranded molecule designde based on mature miR-155,which mimics the high expression status of miR-155 in vivo)and its negative control sequences were transfected with EBV-Ramous,miR-155-mimics(Ramous-mi)、Ramous-yin(Ramous-CON).Contimue to use Sybgreen fluorescence quantitative PCR to detect the expression level of miR-155 in each group of cells after transfection.Western Blot method was used to detect the expression of LMP1(Latent membrane protein1)in each group of cells after transfection.The cell proliferation rate of each group after transfection was detected by CCK8 method.Using SPSS24.0 statistical analysis software to analyze the experimental data,"mean±standard"is used to indicate the measurement data,t test is used to compare the mean between the two groups,P<0.05 is considered statistically significant.Research results:After EBV~+Raji cells and EBV~-Ramous cells were cultured,the expression level of miR-155 was detected by Sybgreen fluorescence quantitative PCR.The relative expression level of miR-155 in EBV~+Raji cells was 5.78±4.15.In EBV~-Ramous The relative expression level in the cells is:0.24±0.14,miR-155 is highly expressed in EBV~+Raji cells,miR-155 is lowly expressed in EBV~-Ramous cells,EBV~+Raji cells are significantly higher than EBV~-Ramous cells,the difference has Statistical significance(P<0.05).Sybgreen fluorescence quantitative PCR was used to detect the expression level of miR-155 in the four groups of cells after transfection.The relative expression level of miR-155 in miR-155-inhibitor(Raji-in)cells was:23.47±1.54;miR-155 The relative expression level in Raji-CON cells is:28.25±0.43;the relative expression level of miR-155 in miR-155-mimics(Ramous-mi)cells is:9461.62±376.98;miR-155 The relative expression in Ramous-CON cells was:158.34±14.48.The expression level of miR-155 in Raji-in is lower than Raji-CON,and the expression level in Ramous-mi is higher than Ramous-CON,the difference is statistically significant(P<0.05).Western Blot method was used to detect the expression of LMP1 in transfected cells.The expression of LMP1 in Raji-in cells was:0.19±0.12,and the expression in Raji-CON cells was:0.37±0.13,in Raji-naked(Untransfected group)The expression level in cells is:0.86±0.16.The expression of LMP1 in Raji-in cells was significantly lower than that of Raji-naked(untransfected group),the difference was statistically significant(P<0.05).Using CCK8 to detect the proliferation rate of the 4 groups of cells at 0 hours,24 hours,48 hours and 72 hours after transfection:the cells in the Raji-in group were:99.32±4.97,92.17±3.32,77.39±3.50,90.16±4.29;Raji-CON The cells in the group were:99.76±6.84,97.43±2.33,62.72±2.14,81.84±3.75;the cells in the Ramous-mi group were:98.94±1.20,78.86±5.08,68.97±6.80,88.55±4.21;the cells in the Ramous-CON group Respectively:97.73±1.92,92.88±2.91,60.29±3.03,81.21±6.40.At 24h,the proliferation rate of Raji-in group was higher than that of Ramous-mi group,and that of Ramous-mi group was lower than that of Ramous-CON group.cell.At 48h,the proliferation rate of Raji-in group was higher than that of Raji-CON group and Ramous-mi group.The cell proliferation rate of Ramous-mi group was higher than that of Ramous-CON group,the difference was statistically significant(P<0.05).Inhibition rates of cells in 4 groups after transfection at 0 hours,24 hours,48 hours,and 72 hours:Raji-in group cells were:0.68±4.97,7.83±3.32,22.61±3.50,9.84±4.29;Raji-CON group cells were It is:0.24±6.84,2.57±2.33,37.28±2.14,18.16±3.75;the cells in the Ramous-mi group are:1.06±1.20,21.14±5.08,31.03±6.80,11.45±4.21;the cells in the Ramous-CON group are:2.67±1.92,7.12±2.91,39.71±3.03,18.79±6.40.At 24h,the inhibition rate of Raji-in group was lower than that of Ramous-mi group,and the inhibition rate of Ramous-mi group was higher than that of Ramous-CON group.There was a significant difference(P<0.05).At 48h,the inhibition rate of Raji-in group was lower than that of Raji-CON group and Ramous-mi group,and the inhibition rate of Ramous-mi group was lower than that of Ramous-CON group,the difference was statistically significant(P<0.05).Conclusion:miR-155 is closely related to many human tumors.It is one of the most widely studied miRNAs in B-cell malignancies and may become a new therapeutic target in the future.In this paper,the culture of Burkitt lymphoma cells and Sybgreen fluorescence quantitative PCR detection showed that miR-155 was significantly overexpressed in EBV~+Raji,which was significantly higher than EBV~-Ramous cells(P<0.05).Western Blot method detected that the expression of LMP1 in Raji-in cells was significantly lower than that of Raji-CON cells and Raji-naked cells(untransfected group)(P<0.05),indicating the inhibition of miR-155 expression in EBV~+Raji cells Will inhibit the expression of LMP1 in cells.CCK8 method was used to detect the inhibition rate of miR-155 and the proliferation rate of cells in each group.At 24h,the proliferation rate of Raji-in group was higher than that of Ramous-mi group,and that of Ramous-mi group was lower than that of Ramous-CON group.(P<0.05),the inhibition rate of Raji-in group was lower than that of Ramous-mi group,and the inhibition rate of Ramous-mi group was higher than that of Ramous-CON group(P<0.05).It indicated that promoting the expression of miR-155 inhibited cell proliferation in EBV~-Ramous cells at 24 hours.At 48h,the cell proliferation rate of Raji-in group was higher than that of Raji-CON group and Ramous-mi group,the cell proliferation rate of Ramous-mi group was higher than that of Ramous-CON group(P<0.05),Raji-in group The cell suppression rate was lower than that in the Raji-CON group and Ramous-mi group,and the cell suppression rate in the Ramous-mi group was lower than that in the Ramous-CON group(P<0.05),indicating that the expression of miR-155 was inhibited in 48 hours of EBV~+Raji cells Promote cell proliferation.These studies provide a data basis for further research on the role of miR-155 in lymphoma cells. |
