The Effect Of α-synuclein On Astrocytes Secreting GDNf And Regulating The Growth Of Neuron Axons

Objective:To investigate the effects of overexpression of a-synuclein WT and mutatants(A30P、A53T)on secretion of GDNF in AST and growth of neurons.Method:AST and neurons were isolated from newborn mice within1-3days and within24hours, resepectively.Endogenous a-synuclein was detected by Western blot in primary cultured AST. After48and72hours transfection the lentivirus expressing wide type a-synuclein and mutatants(A30P、A53T) were obtained after co-transfection of WT,A30P,A53Ta-synuclein-HA-FUIGW-GFP with helper plasmids in293FT cells.Exogenous a-synuclein-HA and mutatants were identified by immunofluorescence and Western blot. The levels of GDNF in conditioned medium of the trasfected AST were measured using enzyme linked immunosorbent assay (ELISA). Transwell system to co-culture the transfected AST and neurons were used.After4days co-culture,length of the neuron axons were measured by immunofluorescence.Result:1. Endogenous a-synuclein was expressed in primary cultured AST.2.7days after the transfection of AST, ectopic a-synuclein (WT) -HA、α-synuclein (A30P)-HA、α-synuclein(A53T)-HA were identified in the transfected AST by immunofluorescence and Western blot.3.The levels of GDNF of WT、A30P、 A53Tα-synuclein groups are less than the FUIGW group in conditioned medium of the trasfected AST (P<0.01).4.4days after co-culturing of AST and neurons,the length of the neuron axons in WT、A30P、A53Tα-synuclein groups are shorter than the FUIGW group (P<0.01)Conclusion:Overexpression of a-synuclein and mutatants(A30P、A53T) decreases the secretion of GDNF and may inhibit the growth of the neuron axons.