Inhibition Effect Of Chemically Synthetic SiRNA On GFAP MRNA In Astrocyte

Objective: Astrocyte is very important in normal nervous activity and the regeneration of central nerve system. glial fibrillary acidic protein(GFAP)which is related with spinal cord injuries is the identification marker of Astrocyte. This research is aim to investigate the inhibition effect of chemically synthetic small interfering RNA (siRNA) on the expression of glial fibrillary acidic protein in astrocyte in rats,and then we select the effect sequence for the next research.Methods: Separate astrocyte from cerebral cortex and culture in DMEM/F12 with 20% FBS, identified by GFAP-FITC in immunofluorescence method. Three pairs of 21 nucleotide(nt) siRNAs-directed rats'GFAP mRNA were differently transfected into through TransMessenger? Transfection Reagent, untreated and transfected nonspecific siRNA astrocyte as the blank control and the nonspecific siRNA control , respectively. The expression of GFAP in mRNA level were assessed by RT-PCR.Results: Three pairs of siRNAs reduced the mRNA level in astrocyte in concentration-dependent manner within 1～3 ug/uL , and the maximum inhibition rate was found in the second siRNA in 2ug/ L ; nonspecific siRNA could not affect GFAP mRNA expression in astrocyte. The expression of GFAP mRNA was reduced by (82±1) %,(87±2) % and (75±4) % (P<0.01), respectively, in siRNA-transfected astrocyte compared with those in blank control .And the expression was decreased by (90±4) %, (64±3) % and (11±6) % (P<0.01) at 24hour,48hour and 72hour after transfection. It show that the effect is time-dependent within 72hours, and the maximum inhibit effect is at 24 hour.