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Comparison of adeno-associated viral 2 vector and lentiviral vector transfection in rat retina after intravitreal injection
HUANG Ting-ting CAO Wen-luo ZHANG Shou-mei Wang Dong XU Jia-jun*
Acta Anatomica Sinica ›› 2016 ›› Issue (2) : 191-196.
Comparison of adeno-associated viral 2 vector and lentiviral vector transfection in rat retina after intravitreal injection
Objective To compare the transfection of recombinant adeno-associated viral 2 vector (rAAV2) with lentiviral vector (LV) in rat retina after intravitreal injection, in order to supply experimental basis of vector choice for gene therapy of ophthalmic diseases and injuries. Methods Twelve rats each group of sixty in the experimental groups were accepted rAAV2-EGFP, rAAV2-neuritin-EGFP, LV-RFP or LV-neuritin-RFP by intravitreal injection respectively, whereas rats in the control group, normal saline. Four weeks later, retinas of all rats were taken for observation. The immunohistochemical staining and CTB-FITC were used to determine the cell types and percentage of transfection. Real-time PCR and Western blotting were used to detect the expression of mRNA and protein of neuritin in the retina. Results The majority of retinal ganglion cells (RGCs) with a percentage of 70% were transfected by rAAV2, whereas LV transfected pigment epithelium and only 30% of RGCs. The expression of neuritin mRNA and protein upregulated 16-flod and 3-fold respectively in the rAAV2-neuritin-EGFP group compared with the rAAV2-EGFP group and the control group, whereas the expression of neuritin mRNA and protein upregulated 5.5-flod and 1.7-fold in the LV-neuritin-RFP group compared with the LV-RFP group and the control group. Conclusion After intravitreal injections, rAAV2 but not LV could provide majority RGCs transduction and more overexpression of neuritin, LV mainly transfect the pigment epithelium. The results suggest that when RGCs need to be transfected, we should choose rAAV2, and when pigment epithelium need to be transfected, we should choose LV in the gene therapy of ophthalmic diseases and injuries.
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