Surgucheva Irina, Weisman Alejandra D, Goldberg Jeffrey L, Shnyra Alexander, Surguchov Andrei
Retinal Biology Research Laboratory, Veterans Administration Medical Center, Kansas City, MO 64128, USA.
Mol Vis. 2008 Aug 22;14:1540-8.
Previous studies have described gamma-synuclein as a protein highly expressed in retinal ganglion cells (RGCs), and a loss of RGCs correlates with a downregulation of gamma-synuclein gene expression in glaucoma. Here we asked whether gamma-synuclein expression in the retina can be considered a specific marker of RGCs.
gamma-Synuclein expression was examined with immunohistochemistry in retinal sections from normal and glaucomatous human eyes. Primary cultures of RGCs from Sprague-Dawley rats purified by sequential immunopanning using a monoclonal antibody to Thy1-1, cultures of A7 immortalized optic nerve astrocytes from newborn rats, and the immortalized RGC-5 cell line were studied using immunofluorescence and quantitative RT-PCR.
gamma-Synuclein was highly expressed in RGCs in the human retina and was localized in cytoplasm adjacent to the RGC nuclear marker, Brn-3a. Axons of RGCs were immunopositive for gamma-synuclein in the nerve fiber layer (NFL), the lamina cribrosa and the retrobulbar optic nerve. In the optic nerve of glaucoma patients, axon swellings were likewise immunopositive, whereas in the retina of patients with retinoblastoma, NFL staining appeared reduced. In primary rat RGCs and in immortalized RGC-5 cultures, gamma-synuclein was localized predominantly in the perinuclear area and in cell processes. Among rat retinal cells in culture, all Brn-3a positive cells were stained with a gamma-synuclein antibody; rare gamma-synuclein-positive cells were not stained by the Brn-3a antibody.
gamma-Synuclein is selectively and abundantly expressed in human RGCs in vivo, primary rat RGCs in vitro, and immortalized RGC-5 cells. In pathology, gamma-synuclein abundance may vary between RGC somas and axons. Coincident Brn-3a and gamma-synuclein expression suggests that strong gamma-synuclein expression can be considered a marker of RGCs. Future translational approaches might include using a gamma-synuclein promoter for the specific delivery of siRNA or therapeutic proteins to RGCs.
以往研究表明γ-突触核蛋白是一种在视网膜神经节细胞（RGCs）中高度表达的蛋白质，在青光眼中RGCs的丧失与γ-突触核蛋白基因表达的下调相关。在此，我们探讨视网膜中γ-突触核蛋白的表达是否可被视为RGCs的特异性标志物。
采用免疫组织化学方法检测正常人和青光眼患者人眼视网膜切片中γ-突触核蛋白的表达。使用针对Thy1-1的单克隆抗体通过连续免疫淘选法纯化的Sprague-Dawley大鼠RGCs原代培养物、新生大鼠的A7永生化视神经星形胶质细胞培养物以及永生化RGC-5细胞系，采用免疫荧光和定量RT-PCR进行研究。
γ-突触核蛋白在人视网膜的RGCs中高度表达，并定位于与RGC核标志物Brn-3a相邻的细胞质中。RGCs的轴突在神经纤维层（NFL）、筛板和球后视神经中对γ-突触核蛋白呈免疫阳性。在青光眼患者的视神经中，轴突肿胀同样呈免疫阳性，而在视网膜母细胞瘤患者的视网膜中，NFL染色似乎减少。在原代大鼠RGCs和永生化RGC-5培养物中，γ-突触核蛋白主要定位于核周区域和细胞突起中。在培养的大鼠视网膜细胞中，所有Brn-3a阳性细胞均被γ-突触核蛋白抗体染色；罕见的γ-突触核蛋白阳性细胞未被Brn-3a抗体染色。
γ-突触核蛋白在体内的人RGCs、体外的原代大鼠RGCs和永生化RGC-5细胞中选择性且大量表达。在病理学中，γ-突触核蛋白的丰度在RGCs胞体和轴突之间可能有所不同。Brn-3a和γ-突触核蛋白的共表达表明，强烈的γ-突触核蛋白表达可被视为RGCs的标志物。未来的转化方法可能包括使用γ-突触核蛋白启动子将siRNA或治疗性蛋白质特异性递送至RGCs。
