Upregulation of EphB2 and ephrin-B2 at the optic nerve head of DBA/2J glaucomatous mice coincides with axon loss
Upregulation of EphB2 and ephrin-B2 at the optic nerve head of DBA/2J glaucomatous mice coincides with axon loss
复制标题
DOI：
10.1167/iovs.07-0442
复制
发表时间：
2007-12-01
影响因子：
4.4
通讯作者：
Sretavan, David W.
中科院分区：
文献类型：
--
作者：
Du, Juan;Tran, Tony;Sretavan, David W.
目的.鉴定与视网膜神经节细胞（RGC）轴突缺失一致的视神经乳头（ONH）表达上调的基因。为了进一步证明这些基因编码的蛋白质与RGC轴突结合并影响基本轴突生理学。对DBA/2 J小鼠（3至11月龄）和C57 Bl/6 NCrl小鼠（10月龄）的ONH切片进行原位杂交和细胞类型特异性免疫标记。将EphB 2-Fc和肝配蛋白-B2-Fc嵌合蛋白在体外和体内应用于ONH的成人RGC轴突，以证明轴突上的蛋白结合。将EphB 2-Fc或对照Fc蛋白在浴中或局部施加到预加载有钙指示剂Fluo-4-AM的轴突上，并测定轴突内钙的变化。EphB 2和肝配蛋白-B2在9月龄开始的DBA/2 J小鼠的ONH处特异性上调，但在年龄匹配的C57 Bl/6 NCrl小鼠或没有轴突损失的DBA/2 J动物中不上调。EphA 4也存在于ONH中，但未受影响的动物和受影响的动物之间没有检测到表达差异。EphB 2由F4/80（+）、MOMA 2（+）、ED 1（-）巨噬细胞样细胞表达，ephrin-B2由Iba-1（+）小胶质细胞和GFAP（+）星形胶质细胞表达，而EphA 4由GFAP（+）星形胶质细胞表达。EphB 2-Fc和肝配蛋白-B2-Fc蛋白在培养物中结合至RGC轴突并且在体内结合至ONH RGC轴突。体外培养的成年RGC轴突对EphB 2-Fc的反应是轴突内钙离子升高，而对对照Fc蛋白无反应。EphB 2和ephrin-B2的表达在与RGC轴突损失一致的胶质瘤DBA/2 J小鼠的ONH处上调。EphB 2和Eph-rin-B2在ONH处的成人RGC轴突上的直接结合以及EphB 2升高轴突内钙的能力表明，这些蛋白质可能在青光眼的情况下影响RGC轴突生理学，从而影响疾病的发展或进展。
PURPOSE. To identify genes with upregulated expression at the optic nerve head (ONH) that coincides with retinal ganglion cell (RGC) axon loss in glaucomatous DBA/2J mice. To further demonstrate that the proteins encoded by these genes bind to RGC axons and influence fundamental axon physiology.METHODS. In situ hybridization and cell-type-specific immunolabeling were performed on ONH sections from DBA/2J mice (3 to 11 months old) and C57Bl/6NCrl mice (10 months old). EphB2-Fc and ephrin-B2-Fc chimeric proteins were applied to adult RGC axons in vitro and in vivo at the ONH to demonstrate protein binding on axons. EphB2-Fc or control Fc protein was applied in a bath or locally to axons preloaded with the calcium indicator Fluo-4-AM, and changes in intra-axonal calcium were determined.RESULTS. EphB2 and ephrin-B2 were specifically upregulated at the ONH of DBA/2J mice starting at 9 months of age, but not in age-matched C57Bl/6NCrl mice or in DBA/2J animals that did not have axon loss. EphA4 was also present at the ONH, but no difference in expression was detected between unaffected and affected animals. EphB2 was expressed by F4/80(+), MOMA2(+), ED1(-) macrophage-like cells, ephrin-B2 was expressed by Iba-1(+) microglia and GFAP(+) astrocytes, whereas EphA4 was expressed by GFAP(+) astrocytes. EphB2-Fc and ephrin-B2-Fc protein bound to RGC axons in culture and to ONH RGC axons in vivo. Adult RGC axons in vitro elevated intra-axonal calcium in response to EphB2-Fc but not to control Fc protein.CONCLUSIONS. The expression of EphB2 and ephrin-B2 is upregulated at the ONH of glaucomatous DBA/2J mice coinciding with RGC axon loss. The direct binding of EphB2 and Eph-rin-B2 on adult RGC axons at the ONH and the ability of EphB2 to elevate intra-axonal calcium indicate that these proteins may affect RGC axon physiology in the setting of glaucoma and thus affect the development or progression of the disease.