Effect On Biological Property And Osteoblastic Differentiation Of MG63 Cells By N-acetyl-L-Leucine-modified Polyethyleniminemediated MiR-34a Delivery

Objective:To transfect the N-Ac-L-Leu-PEI mediated miR-34 a into the MG63 cells,and to explore the characterization of N-Ac-L-Leu-PEI /miR-34 a complex,and the effect on biological property and and osteoblastic differentiation in MG63 cells by N-Ac-L-Leu-PEI /miR-34 a complex.Methods:Firstly,in order to screen out the optimal transfection quality ratio of N-Ac-L-Leu-PEI/miR-34 a,The N-Ac-L-Leu-PEI was blended with miR-34 a by different mass ration.Using the Malvin particle potential to test the particle size and potential of N-Ac-L-Leu-PEI / miR-34 a complex and by agarose gel electrophoresis to determinate the ability of N-Ac-L-Leu-PEI with miR-34 a.The transfection efficiency and m RNA expression of miR-34 a in MG63 cells with different mass ration were detected by fluorescence imaging technique,flow cytometry and RT-PCR,which was to determine the optiomal transfection mass ratio.Secondly,to evaluate the effect of N-Ac-L-Leu-PEI/miR-34 a on the biological properties of MG63 cells,the effect of MTT assay on the proliferation of MG63 cells was investigated.Determined the cell cycle and apoptosis of N-Ac-L-Leu-PEI / miR-34 a complex by flow cytometry.Lastly,RT-PCR and Western blot were used to detect the changes of the expression of osteoblast-related genes in MG63 cells,and to explore the effect of N-Ac-L-Leu-PEI/miR-34 a on osteoblast differentiation.Results:N-Ac-L-Leu-PEI/miR-34 a complex with the increase of mass ratio,the particle size decreases,the potential increased,and the mass ratio of greater than or equal to 2:1,miR-34 a was fully loaded by N-Ac-L-Leu-PEI to form a stable complex.What is more,when the mass ratio was 4: 1,the transfection efficiency of N-Ac-L-Leu-PEI / miR-34 a was the highest in MG63 cells.Compared with the blank control group,N-Ac-L-Leu-PEI / miR-34 a complex inhibited cell proliferation,inhibited cell cycle,and had no significant effect on cell apoptosis.The results of RT-PCR and Western blot analysis showed that the expression of Runx2,SP7 and Col I genes was promoted by N-Ac-L-Leu-PEI / miR-34 a group,compared with the blank control group.Conclusion:The miR-34 a can be effectively transfected into MG63 cells and was highly expressed in the cells by using N-Ac-L-Leu-PEI as the vector.Mi R-34 a has a certain ability of inhibiting the proliferation and promoting osteogenesis differentiation of MG63 cells.