Th E Effect Of MiR-328 On The Proliferation Of K562 Cells And Its Mechanism

Aim:Construction of miR-328 over expression and down-regulation vector. Our research is aim to investigate the effect of miR-328 on proliferation of chronic myeloid leukemia(CML) blast crisis cell K562, and the mechanism. Methods:Construction of mi R-328 overexpression and down-regulation vector: hsa-miR-328 and hsa-miR-328-inhibition, recombinant plasmid by nucleotide sequencing identification. Recombinant plasmid transfected into K562 cells respectively by lipofectamineTM2000. Oberve the transfected cell under Fluorescence microscope. Detect the expression level of miR-328 and C/EBP α mRNA by real-time fluorescence quantitative PCR; CCK-8 was used to estimating the cell viability. Determination of C/EBP α protein expression by use Western-Blot method; Results:1. Our study was successfully constructed vector : hsa-mi R-328 and hsa-miR-328-inhibition;2. Plasmid vectors were transfected into K562 cells. Fluorescent cells were observed after 24 h, and the visible fluorescence cells were gradually increased after 48 h or 72 h.3. Real time fluorescence quantitative PCR results showed that compared with control group、Lip2000 group、empty vector group、hasmiR-328-inhibition group, hsa-miR-328 group miR-328 showed obvious expression, the difference was significant(P < 0.001);.4. The results of CCK-8 showed that compared with control group、Lip2000 group、empty vector group、has-miR-328-inhibition group, hasmiR-328 group the proliferat-ion of K562 cells was significantly inhibited, the difference was significant(P < 0.001);5. The results of real-time quantitative PCR showed no significant difference in the expression of C/EBP alpha mRNA in K562 cells, compared with control group、Lip2000 group、empty vector group、has-miR-328-inhibition group(P > 0.05);6. Western-Blot detected that the expression of C/EBP protein in K562 cells of has-miR-328 group, C/EBP protein was not expressed or low level in other groups. Conclusion:miR-328 inhibits the proliferation of K562 cells.the mechanisim by upregulation of C/EBP α.