关键词: 星形胶质细胞, 视神经再生, 青光眼, 转基因小鼠

Abstract: Objective To summarize techniques and experience in developing mice optic nerve crushing model for studying optic nerve regeneration.Methods 4 Bcl-2/GFAP-Tk double transgenic mice were obtained by mating male Bcl-2 transgenic mice with female mice expressing herpes simplex virus-thymidine kinase transgenic mice controlled under GFAP promoter(GFAP-Tk) and 4 Bcl-2 transgenic mice were used as controls,and ganciclovir was administered through slow-releasing osmotic pump to selectively eliminate activated astrocytes in Bcl-2/GFAP-Tk transgenic mice.The right optic nerve crush was carded out according to the standard procedure in these mice aged 8～12 weeks(20～30g) on the second day after implantation of the osmotic pump.The tissue samples were taken on the tenth day after the optic nerve crush,and the axon regeneration was analyzed qualitatively and quantitatively by GAP43 immunofluorescence staining.Rodamine conjugate CTB or replication defective adeno associate virus carrying enhanced green fluorescence protein(AAV-EGFP) was injected in vitreous to label axon anterogradely and to determine whether the regenerated axons reach the brain targets.Results The remarkable optic nerve regeneration and the growth cone-like structure were found and the regenerated axon count was 71.99±24.04 in Bcl-2/GFAP-Tk double transgenic mice after GCV administration and optic nerve crushing,but the axon reached into the brain was found.No axonal regeneration was identified in Bcl-2 transgenic mice.Conclusion Complete optic nerve crushing in the mice is an ideal mode for studying neural regeneration,but stable techniques of developing the mode and specific labeling methods for axonal regeneration are imperative.

Key words: Astrocyte, Optic nerve regeneration, Glaucoma, Transgenic mice