〔Abstract〕 Objective Eya1 gene lentiviral vectors were constructed, and the effects of overexpression of eye absent 1(Eya1) on 6-OHDA-induced apoptosis of MES23.5 dopaminergic cells were investigated. Methods Lentivirus overexpression of Eya1 gene was constructed and infected MES23.5 cells, then stable Eya1-expression cell lines were screened. MES23.5 dopaminergic cell injury model induced by 6-hydroxydopamine(6-OHDA) was divided into control group, empty vector group and overexpression Eya1 group. Cell counting kit-8(CCK-8), Hoechst staining kit(Hoechst 33258) and Western blot were used to detect the effect of overexpression of Eya1 on apoptosis. Results Enzyme electrophoresis showed that the target gene was recombined into lentivirus vector. Sequencing results showed that Eya1 gene overexpression vector was identical with the Eya1 gene coding sequence. CCK-8 showed that the cell viability of Eya1 overexpressed group was higher than that of control group(P<0.05). Hoechst 33258 showed that the concentration and aggregation level of Eya1 overexpression group was lower than that of control group. The expression of Bcl-2 protein in the overexpression Eya1 group was higher than that in the control group(P<0.05), while that of Bax protein was lower than that of the control. Conclusion The MES23.5 cell line overexpressing of Eya1 was successfully constructed. Eya1 antagonizes the apoptosis of MES23.5 dopaminergic cells induced by 6-OHDA by regulating the expression of Bcl-2 and Bax proteins.