Gene therapy with brain-derived neurotrophic factor as a protection: Retinal ganglion cells in a rat glaucoma model
Gene therapy with brain-derived neurotrophic factor as a protection: Retinal ganglion cells in a rat glaucoma model
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DOI：
10.1167/iovs.02-1332
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发表时间：
2003-10-01
影响因子：
4.4
通讯作者：
Hauswirth, WW
中科院分区：
文献类型：
--
作者：
Martin, KRG;Quigley, HA;Hauswirth, WW
目的.目的：构建一种能够有效转染视网膜神经节细胞（RGCs）的腺相关病毒（AAV）载体，并验证该载体表达脑源性神经营养因子（BDNF）对实验性胶质瘤的保护作用。93只大鼠接受一次单侧玻璃体内注射生理盐水（n = 30）、AAV-BDNF-土拨鼠肝炎转录后调节元件（WPRE; n = 30）或AAV-绿色荧光蛋白（GFP）-WPRE（n = 33）。两周后，通过对小梁网施加激光在注射眼中诱导实验性青光眼。通过计算青光眼4周后视神经横截面中的轴突来估计RGCs的存活。通过免疫组织化学、蛋白质印迹分析和直接可视化GFP.视网膜整体切片中GFP阳性细胞的密度为1，828 +/- 299个细胞/mm（2）（72，273 +/-11，814个细胞/视网膜）。所有组中眼内压升高的暴露相似。初始激光治疗后4周，盐水治疗组（n = 25）和AAV-GFP-WPRE组（n = 30）的轴突损失分别为52.3% +/- 27.1%和52.3% +/- 24.2%，但AAV-BDNF-WPRE组（n = 27）仅为32.3% +/- 23.0%。AAV-BDNF-WPRE动物的存活率显著增加，与接受AAV-GFP-WPRE（P = 0.002，t检验）或盐水（P = 0.006，t检验）的动物相比，差异显著。在大鼠青光眼模型中通过轴突计数估计，BDNF基因的过表达保护RGC，进一步支持神经营养疗法作为青光眼治疗中降低IOP的补充的潜在可行性。
PURPOSE. To develop a modified adenoassociated viral (AAV) vector capable of efficient transfection of retinal ganglion cells (RGCs) and to test the hypothesis that use of this vector to express brain-derived neurotrophic factor (BDNF) could be protective in experimental glaucoma.METHODS. Ninety-three rats received one unilateral, intravitreal injection of either normal saline (n = 30), AAV-BDNF-woodchuck hepatitis posttranscriptional regulatory element (WPRE; n = 30), or AAV-green fluorescent protein (GFP)-WPRE (n = 33). Two weeks later, experimental glaucoma was induced in the injected eye by laser application to the trabecular meshwork. Survival of RGCs was estimated by counting axons in optic nerve cross sections after 4 weeks of glaucoma. Transgene expression was assessed by immunohistochemistry, Western blot analysis, and direct visualization of GFP.RESULTS. The density of GFP-positive cells in retinal whole-mounts was 1,828 +/- 299 cells/mm(2) (72,273 +/- 11,814 cells/ retina). Exposure to elevated intraocular pressure was similar in all groups. Four weeks after initial laser treatment, axon loss was 52.3% +/- 27.1% in the saline-treated group (n = 25) and 52.3% +/- 24.2% in the AAV-GFP-WPRE group (n = 30), but only 32.3% +/- 23.0% in the AAV-BDNF-WPRE group (n = 27). Survival in AAV-BDNF-WPRE animals increased markedly and the difference was significant compared with those receiving either AAV-GFP-WPRE (P = 0.002, t-test) or saline (P = 0.006, t-test).CONCLUSIONS. Overexpression of the BDNF gene protects RGC as estimated by axon counts in a rat glaucoma model, further supporting the potential feasibility of neurotrophic therapy as a complement to the lowering of IOP in the treatment of glaucoma.