AAV-mediated expression of CNTF promotes long-term survival and regeneration of adult rat retinal ganglion cells
AAV-mediated expression of CNTF promotes long-term survival and regeneration of adult rat retinal ganglion cells
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DOI：
10.1038/sj.gt.3302791
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发表时间：
2006-09-01
期刊：
影响因子：
5.1
通讯作者：
Harvey, A. R.
中科院分区：
文献类型：
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作者：
Leaver, S. G.;Cui, Q.;Harvey, A. R.
我们比较了玻璃体内注射编码增强型绿色荧光蛋白（GFP）和睫状神经营养因子（CNTF），脑源性神经营养因子（BDNF）或生长相关蛋白-43（GAP 43）对（i）视神经（ON）挤压或（ii）后成人视网膜神经节细胞（RGC）存活和再生的影响在ON切割和周围神经（PN）的附着之后。在ON挤压后7周，β III-微管蛋白免疫染色的定量显示，与AAV-GFP对照相比，AAV-GAP 43-GFP没有增强RGC存活，但AAV-CNTF-GFP增加了RGC存活（平均RGC/视网膜：17450 +/-358 s.e.m.）和AAV-BDNF-GFP注射的眼睛（10200 +/-4064个RGC/视网膜）。与AAV-CNTF-GFP和AAV-BDNF-GFP注射的眼睛中增加的RGC活力一致，这些动物在ON挤压近端具有许多β III-微管蛋白和GFP阳性纤维。然而，仅在AAV-CNTF-GFP组中，在远端ON中观察到再生的RGC轴突（11357367个轴突/神经，挤压后0.5mm），一些到达视交叉。RGC对CNTF呈免疫反应性，定量RT-PCR显示用AAV-CNTF-GFP转导的视网膜中CNTF mRNA表达显著增加。RGC的AAV-CNTF-GFP转导与自体PN-ON移植的组合导致甚至更大的RGC存活和再生。PN移植后7周，视网膜上存活的RGC为27954（72833）个，约占成年RGC总数的25%。其中，13352（71868）个RGCs/视网膜在PN移植物中注射荧光金后被逆行标记。总之，AAV介导的CNTF表达促进损伤的成年RGC的长期存活和再生，通过组合基于基因和细胞的治疗/干预显著增强了这种作用。
We compared the effects of intravitreal injection of bicistronic adeno-associated viral (AAV-2) vectors encoding enhanced green fluorescent protein (GFP) and either ciliary neurotrophic factor ( CNTF), brain-derived neurotrophic factor ( BDNF) or growth-associated protein-43 (GAP43) on adult retinal ganglion cell (RGC) survival and regeneration following (i) optic nerve ( ON) crush or (ii) after ON cut and attachment of a peripheral nerve (PN). At 7 weeks after ON crush, quantification of beta III-tubulin immunostaining revealed that, compared to AAV-GFP controls, RGC survival was not enhanced by AAV-GAP43-GFP but was increased in AAV-CNTF-GFP ( mean RGCs/retina: 17450 +/- 358 s.e.m.) and AAV-BDNF-GFP injected eyes (10200 +/- 4064 RGCs/retina). Consistent with increased RGC viability in AAV-CNTF-GFP and AAV-BDNF-GFP injected eyes, these animals possessed many beta III-tubulin- and GFP-positive fibres proximal to the ON crush. However, only in the AAV-CNTF-GFP group were regenerating RGC axons seen in distal ON ( 11357367 axons/nerve, 0.5mm post-crush), some reaching the optic chiasm. RGCs were immunoreactive for CNTF and quantitative RT-PCR revealed a substantial increase in CNTF mRNA expression in retinas transduced with AAV-CNTF-GFP. The combination of AAV-CNTF-GFP transduction of RGCs with autologous PN-ON transplantation resulted in even greater RGC survival and regeneration. At 7 weeks after PN transplantation there were 27 954 ( 72833) surviving RGCs/retina, about 25% of the adult RGC population. Of these, 13 352 (71868) RGCs/retina were retrogradely labelled after fluorogold injections into PN grafts. In summary, AAV-mediated expression of CNTF promotes long-term survival and regeneration of injured adult RGCs, effects that are substantially enhanced by combining gene and cell-based therapies/interventions.