*Centre for Ophthalmology, University Eye Hospital Tübingen, Tübingen †Department of Ophthalmology, Charite Berlin, Berlin, Germany.
J Glaucoma. 2014 Apr-May;23(4):219-24. doi: 10.1097/IJG.0000000000000040.
Cyclosporine A (CSA) is a widely used immunosuppressive drug. Furthermore, CSA showed neuroprotective properties in several neurological disorders. However, nearly no data exist regarding CSA and its possible neuroprotective effect on retinal ganglion cells (RGCs).
RGC-5 cells were stressed with 10 mM glutamate for 24 hours with or without adding varying concentrations of CSA (1, 3, 6, or 9 μg/mL) to the medium. Cell viability and cell density were analyzed by MTS assay and crystal violet staining, respectively. Induction of apoptosis was determined through caspase 3/7 activity and Annexin V+/PI- flow cytometry.
The incubation of RGC-5 cells with 10 mM glutamate for 24 hours induced a 3.1-fold and a 3.4-fold decrease in overall cell viability and cell density, respectively, compared with controls. The supplementation of 9 μg/mL CSA to 10 mM glutamate led to a 2.7-fold increase in overall cell viability (P<0.0005) and a 2.5-fold increase in cell density (P<0.0005) compared with RGC-5 cells treated only with 10 mM glutamate. Furthermore, the addition of 9 μg/mL CSA to 10 mM glutamate significantly reduced caspase 3/7 activity by 1.3-fold (P<0.0005) and the amount of Annexin V+/PI- cells by 2.8-fold compared with RGC-5 cells incubated with 10 mM glutamate alone. The neuroprotective effect of CSA dose-dependently decreased with lower concentrations.
CSA can effectively protect RGC-5 cells against glutamate-induced excitotoxicity. Therefore, CSA should be tested in further experiments to evaluate its potential as a neuroprotective substance against RGC disorders.
环孢素 A（CSA）是一种广泛应用的免疫抑制剂。此外，CSA 在多种神经疾病中表现出神经保护作用。然而，几乎没有关于 CSA 及其对视网膜神经节细胞（RGC）可能的神经保护作用的数据。
用 10mM 谷氨酸处理 RGC-5 细胞 24 小时，或在培养基中加入不同浓度的 CSA（1、3、6 或 9μg/mL）。通过 MTS 测定法和结晶紫染色分别分析细胞活力和细胞密度。通过 caspase 3/7 活性和 Annexin V+/PI-流式细胞术测定细胞凋亡的诱导。
与对照组相比，用 10mM 谷氨酸孵育 RGC-5 细胞 24 小时分别导致总细胞活力和细胞密度降低 3.1 倍和 3.4 倍。与仅用 10mM 谷氨酸处理的 RGC-5 细胞相比，向 10mM 谷氨酸中添加 9μg/mL CSA 导致总细胞活力增加 2.7 倍（P<0.0005），细胞密度增加 2.5 倍（P<0.0005）。此外，与仅用 10mM 谷氨酸孵育的 RGC-5 细胞相比，向 10mM 谷氨酸中添加 9μg/mL CSA 使 caspase 3/7 活性降低 1.3 倍（P<0.0005）， Annexin V+/PI-细胞数量减少 2.8 倍。CSA 的神经保护作用呈剂量依赖性降低，低浓度时效果下降。
CSA 可有效保护 RGC-5 细胞免受谷氨酸诱导的兴奋性毒性。因此，应在进一步的实验中测试 CSA，以评估其作为治疗 RGC 疾病的神经保护物质的潜力。
