Gap junctions enhance the antiproliferative effect by transfer of microRNAs in glioma cells / 中国药理学与毒理学杂志

ABSTRACT

OBJECTIVE To investigate the permeability of gap junction composed of connexin 43 (Cx43) for microRNAs (miRNAs) and the impact of gap junction-mediated transfer of miRNAs in glioma U87 cells. METHODS Co-culture assay demonstrated the transmission of miRNAs through gap junction channel into adjacent cells. U87 cells were labeled with green fluorescein protein (GFP) as receivers and cells were transfected miRNAs as donors. Receiver cells and donor cells were mixed together in a ratio of 11. After 12 h co-culture, cells were separated using a BD influx flow cytometer based on the GFP labeled. Quantitative real-time polymerase chain reaction (qRT-PCR) was applied detect to the expressions of miRNAs and Cx43 mRNA. Western blotting was performed to detect the protein expressions of Cx43 and GFP in U87 cells. CCK-8 assay is used to detect cell growth. RESULTS Co-culture assays demonstrated miR-34a could transfer between U87 cells. The role of the contact independent could also transfer of miR-34a. Gap junctions inhibitor (CBX and 18-α-GA) showed lower miR-34a expression than co-culture group, whereas gap junctions enhancer (RA and Galanglin) enhanced miR-34a expression. Knockdown of Cx43 could significantly decrease the transferring of miR-34a between U87 cells. Different length of miRNAs (miR-1827, miR-144, miR-203a and miR-1183) were similar to the expression of miR-34a between U87 cells. Additionally, we demonstrated that gap junctions mediate the effect of antiproliferation mediated by miR- 34a in U87 cells. The functional inhibition of gap junctions using either siRNA or inhibition eliminated the miR- 34a mediated antiproliferation, whereas the enhancement of gap junctions treatment augmented this miR34a-mediated antiproliferation. CONCLUSION Our study demonstrates that gap junction composed of Cx43-mediated transfer miRNAs in different length of nucleotides and gap junction-mediated transfer of miR-34a enhance the antiproliferative effect in glioma U87 cells.