Effects of hBDNF-GFP gene-transfected neural stem cell transplantation on BDNF expression in the retina of rats following optic nerve crush injury

Abstract

Objective To investigate the effects of human Rrain-derived neurotrophic factor-gamma fetopro-tein (hBDNF-GFP) gene-transfected neural stem cell (NSC) transplantation on BDNF expressions in the retina of rots after optic nerve (ON) crush injury. Method ①Seventy-eight Sprague-Dawley (SD) rats were randomly as-signed into a control group (n = 6) and ON crush group (n = 72). In the ON crush group, the right ON was crushed while the left NO was exposed as sham injury. Rats in the ON crush group were divided into three sub-groups: PBS group (intravitreons injection of 0.01 mol/L phosphate buffered solution); GFP group (intravitreous transplantation of GFP gene-transfected NSCs); and hBDNF-GFP group (intravitreous transplantation of hBDNF-GFP gene-transfected NSCs). Rats were sacrificed 3, 7, 14 and 28 days after transplantation, and BDNF expres-sions in retinal homogenates was detected by using enzyme-linked immunosorbent assay (ELISA). ②The hBDNF-GFP-NSCs were transplanted intravitreous into six rats after ON crush injury. Following this, two rats were sacri-riced 2, 4 and 8 weeks after transplantation. The survival and location of NSCs in host retina were observed by frozen section analysis. ③Adult SD rats were randomly divided into four groups: control group (n = 5); NSC group (NSC transplantation, n = 10); GFP-NSC group (GFP-NSC transplantation, n = 10); and hBDNF-GFP-NSC group (hBDNF-GFP-NSC transplantation, n = 10). Four and eight weeks after transplantation, five rats from every group were sacrificed. Western blot analysis was used to determine retinal BDNF expression. Results ① There was no significant difference in BDNF expression between the control group and sham-injury groups (P >0.05). Three days after NSC transplantation, BDNF expression increased significantly in the three injured sub-groups compared with the sham-injury group, (P 0.05). Seven days after transplantation, there was a significant difference in BDNF expression between the GFP-NSC group and the sham-injury groups (P 0.05). Fourteen and 28 days after transplantation, BDNF expressions decreased in the PBS group and the GFP-NSC groups, while BDNF expressions in the hBDNF-GFP-NSC group increased significant-ly compared with the other three groups (P < 0.05); ②Frozen section showed that transplanted hBDNF-GFP-NSCs could survive and gradually extended to all layers of the host retina. ③Westem blot revealed there were no differences in BDNF expressions between 4-week and 8-week intervals in the hBDNF-GFP-NSC group. Compared with other three groups, BDNF expressions in the retina increased significantly after hBDNF-GFP-NSC transplanta-tion. Conclusions The hBDNF-GFP gene-trausfected NSCs can survive in the host retina and BDNF expressions are stable at a high level. Key words: Brain-derived neurotrophic factor; Stem cell; Transfection; Retina