Quantitative analysis in neurotrophic factor genes of spinal cord with complementary DNA microarray

慢性机械压迫脊髓会造成神经组织损伤，包括损伤平面周围的前角细胞丢失。外源性神经营养因子对神经细胞的传递可以为机械损伤的脊髓提供神经保护作用。研究了携带神经营养素-3(NT-3)基因的腺病毒载体逆行基因导入慢性和进行性机械压迫的小鼠脊髓前角神经元的效果。ADV-NT-3用于通过胸锁乳突肌逆行将ADV-NT-3递送到16周龄、脊髓受压相对较轻的成年小鼠的颈椎副运动神经元。注射AdV-NT-3或AdV-β-半乳糖苷酶基因(LacZ)作为标记基因后4周，对受压颈脊髓进行组织学、免疫组织学和免疫印迹分析。与注射Adv-LacZ的小鼠相比，注射Adv-NT-3的小鼠对NT-3的免疫反应性显著增强。在受压最大的脊髓水平，Nissl、胆碱乙酰转移酶(ChAT)、trkC染色和麦芽凝集素-辣根过氧化物酶(WGA-HRP)标记的前角神经元数量显著高于AdV-LacZ转染组。逆行将NT-3基因转移到小鼠前角神经元，可增加WGA-HRP标记神经元的轴突长度和树枝状突起。我们的结果表明，在完整的动物中，定向逆行传递NT-3基因是可行的，即使在脊髓受到慢性机械压迫的情况下，它也可以提高神经元的存活率。(C)2008年Wiley-Liss，Inc.

Chronic mechanical compression of the spinal cord causes neural tissue damage, including loss of anterior horn cells around the level of injury. Exogenous delivery of neurotrophins to neuronal cells could provide neuroprotection to a spinal cord subjected to mechanical injury. We investigated the efficacy of retrograde gene delivery of adenoviral vector (AdV) carrying neurotrophin-3 (NT-3) gene into twy (twy/twy) mouse spinal cord anterior horn neurons with chronic and progressive mechanical compression at C1-C2 level. AdV-NT-3 was used for retrograde delivery via the sternomastoid muscle to the cervical spinal accessory motoneurons in 16-week-old adult twy mice with relatively mild spinal cord compression. Four weeks after the AdV-NT-3 or AdV-beta-galactosidase cDNA (LacZ) as a marker gene injection, the compressed cervical spinal cord was examined histologically, immunohistologically, and by immunoblot analysis. Immunoreactivity to NT-3 was significantly enhanced in the AdV-NT-3-injected twy mice compared with the AdV-LacZ-injected mice. The numbers of anterior horn neurons of Nissl-, choline acetyltransferase (ChAT)-, and trkC-stained and wheat germ agglutinin-horseradish peroxidase(WGA-HRP)-labeled neurons at the spinal cord level with maximum compression were significantly higher in AdV-NT-3-transfected than in AdV-LacZ-transfected twy mice. Retrograde NT-3 gene transfer to twy mouse anterior horn neurons increased neurite axonal length and arborization of WGA-HRP-labeled neurons. Our results suggest that targeted retrograde NT-3 gene delivery is feasible in the intact animal and that it enhances neuronal survival even under chronic mechanical compression of the spinal cord. (c) 2008Wiley-Liss, Inc.