Department of Ophthalmology, Saga University Faculty of Medicine, Saga, Japan.
Curr Eye Res. 2012 Oct;37(10):907-13. doi: 10.3109/02713683.2012.688161. Epub 2012 May 29.
To investigate neuroprotective effects of siRNA targeted to caspase-3 against ischemia and reperfusion (I/R) injury in rat eyes.
Retinal ischemia was induced in Wistar rats by increasing the intraocular pressure (IOP) to 110 mmHg for 120 min. To examine the effect of siRNA on rat caspase-3, siRNA was injected into the vitreous cavity 24 h prior to induction of retinal ischemia. Eyes were removed at 2, 7 or 14 days later, and then analyzed for the number of retinal ganglion cells (RGCs), the retinal thickness and the amount of apoptosis of the retinal neural cells (as demonstrated by the TUNEL assay). The amount of caspase-3 mRNA was analyzed by rt-PCR. Differences between groups were evaluated by an unpaired t test.
The numbers of RGCs in the saline and non-silencing siRNA controls were reduced significantly at 2 and 7 days after the I/R injury. RGCs were significantly retained in eyes pretreated with siRNA targeted to caspase-3 as compared to the control eyes at 2 days after the I/R injury. Inner retinal thickness in the control eyes was significantly thinner as compared to the treated eyes at 2 and 7 days after the I/R injury. After siRNA treatment, the amount of caspase-3 mRNA was significantly lower when compared to the saline control group.
The injection of siRNA targeted to caspase-3 into the vitreous cavity of rat eyes may block caspase-3, and may thus be able to prevent retinal cell death associated with ischemic injury. As inhibition of the apoptosis pathway may provide a neuroprotective effect, examination of new strategies for treating these disorders needs to be undertaken.
研究针对半胱氨酸天冬氨酸蛋白酶-3 的 siRNA 对大鼠眼缺血再灌注（I/R）损伤的神经保护作用。
通过将眼内压（IOP）升高至 110mmHg 持续 120min 诱导 Wistar 大鼠视网膜缺血。为了研究 siRNA 对大鼠半胱氨酸天冬氨酸蛋白酶-3 的作用，在诱导视网膜缺血前 24h 将 siRNA 注入玻璃体腔。在之后的 2、7 或 14 天取出眼睛，通过 TUNEL 检测分析视网膜神经细胞的凋亡数量、视网膜节细胞（RGC）数量和视网膜厚度。通过 rt-PCR 分析半胱氨酸天冬氨酸蛋白酶-3mRNA 的量。通过独立样本 t 检验评估组间差异。
在 I/R 损伤后 2 和 7 天，生理盐水和非沉默 siRNA 对照组的 RGC 数量明显减少。与 I/R 损伤后 2 天的对照眼相比，用靶向半胱氨酸天冬氨酸蛋白酶-3 的 siRNA 预处理的眼睛中 RGC 保留明显更多。在 I/R 损伤后 2 和 7 天，对照组的内视网膜厚度明显比治疗组薄。与生理盐水对照组相比，siRNA 处理后半胱氨酸天冬氨酸蛋白酶-3mRNA 的量明显降低。
将靶向半胱氨酸天冬氨酸蛋白酶-3 的 siRNA 注入大鼠眼玻璃体腔可能会阻断半胱氨酸天冬氨酸蛋白酶-3，从而可能防止与缺血损伤相关的视网膜细胞死亡。由于抑制凋亡途径可能提供神经保护作用，因此需要对治疗这些疾病的新策略进行研究。
