Neuroscience and Ophthalmology, Institute of Inflammation and Ageing, College of Medical and Dental Sciences, University of Birmingham, Birmingham, B15 2TT, UK.
Cell Death Dis. 2019 Feb 4;10(2):102. doi: 10.1038/s41419-019-1379-6.
Retinal ganglion cells (RGCs) undergo rapid cell death by apoptosis after injury but can be rescued by suppression of caspase-2 (CASP2) using an siRNA to CASP2 (siCASP2). Pigment epithelium-derived factor (PEDF), has neuroprotective and anti-angiogenic functions and protects RGC from death. The purpose of this study was to investigate if suppression of CASP2 is a possible mechanism of neuroprotection by PEDF in RGC. Adult rat retinal cells were treated in vitro with sub-optimal and optimal concentrations of siCASP2 and PEDF and levels of CASP2 mRNA and RGC survival were then quantified. Optic nerve crush (ONC) injury followed by intravitreal injections of siCASP2 or PEDF and eye drops of PEDF-34 were also used to determine CASP2 mRNA and protein reduction. Results showed that PEDF and PEDF-34 significantly suppressed CASP2 mRNA in culture, by 1.85- and 3.04-fold, respectively, and increased RGC survival by 63.2 ± 3.8% and 81.9 ± 6.6%, respectively compared to cells grown in Neurobasal-A alone. RGC survival was significantly reduced in glial proliferation inhibited and purified RGC cultures suggesting that some of the effects of PEDF were glia-mediated. In addition, intravitreal injection of PEDF and eye drops of PEDF-34 after ONC also suppressed CASP2 mRNA levels by 1.82- and 3.89-fold and cleaved caspase-2 (C-CASP2) protein levels by 4.98- and 8.93-fold compared to ONC + PBS vehicle groups, respectively, without affecting other executioner caspases. Treatment of retinal cultures with PEDF and PEDF-34 promoted the secretion of neurotrophic factors (NTF) into the culture media, of which brain-derived neurotrophic factor (BDNF) caused the greatest reduction in CASP2 mRNA and C-CASP2 protein. The neuroprotective effects of PEDF were blocked by a polyclonal antibody and PEDF suppressed key elements in the apoptotic pathway. In conclusion, this study shows that some of the RGC neuroprotective effects of PEDF is regulated through suppression of CASP2 and downstream apoptotic signalling molecules.
视网膜神经节细胞 (RGC) 在受伤后通过细胞凋亡迅速死亡，但可以通过使用 caspase-2 (CASP2) 的 siRNA (siCASP2) 抑制 CASP2 来挽救。色素上皮衍生因子 (PEDF) 具有神经保护和抗血管生成作用，可防止 RGC 死亡。本研究旨在探讨 PEDF 是否通过抑制 CASP2 发挥其对 RGC 的神经保护作用。将成年大鼠视网膜细胞在体外用亚最佳和最佳浓度的 siCASP2 和 PEDF 处理，然后定量检测 CASP2 mRNA 和 RGC 存活率。还使用视神经挤压 (ONC) 损伤后玻璃体内注射 siCASP2 或 PEDF 以及 PEDF-34 滴眼来确定 CASP2 mRNA 和蛋白减少。结果表明，PEDF 和 PEDF-34 在培养物中分别显著抑制了 CASP2 mRNA，抑制率分别为 1.85 倍和 3.04 倍，与仅在 Neurobasal-A 中生长的细胞相比，RGC 存活率分别增加了 63.2%±3.8%和 81.9%±6.6%。在胶质细胞增殖抑制和纯化的 RGC 培养物中，RGC 存活率显著降低，提示 PEDF 的一些作用是通过胶质细胞介导的。此外，ONC 后玻璃体内注射 PEDF 和 PEDF-34 滴眼也分别抑制了 CASP2 mRNA 水平，抑制率分别为 1.82 倍和 3.89 倍，以及 cleaved caspase-2 (C-CASP2) 蛋白水平，抑制率分别为 4.98 倍和 8.93 倍，与 ONC+PBS 载体组相比，而对其他执行 caspase 没有影响。PEDF 和 PEDF-34 处理视网膜培养物可促进神经营养因子 (NTF) 分泌到培养基中，其中脑源性神经营养因子 (BDNF) 导致 CASP2 mRNA 和 C-CASP2 蛋白水平降低最大。PEDF 的神经保护作用被多克隆抗体阻断，PEDF 抑制了凋亡途径中的关键分子。综上所述，本研究表明 PEDF 对 RGC 的部分神经保护作用是通过抑制 CASP2 和下游凋亡信号分子来调节的。
