小干扰RNA保护大鼠视网膜神经节细胞的实验研究

 目的 通过大鼠视神经夹伤模型，研究小干扰RNA（siRNA）对视网膜神经节细胞（RGC）的保护作用。设计 实验研究。 研究对象 SPF级SD大鼠54只。方法 54只SD大鼠随机分为A、B、C三组，每组18只。均选取右眼为实验眼，左眼为正常对照。在球后2 mm处用40 g压力微型视神经夹夹持视神经60 s，做视神经夹伤模型。建立模型后当天，A、B、C三组分别给予玻璃体注射10 μg、20 μg siRNA和生理盐水。视神经夹伤后10天，每组取6只大鼠用荧光金做逆行标记，14天时取标记后的大鼠双眼眼球标本做视网膜铺片并拍摄照片，RGC计数。计算RGC存活率（右眼RGC数/左眼RGC数×100%）。每组其余12只大鼠进一步用蛋白印迹法检测视网膜组织中caspase-3蛋白的表达水平。主要指标 RGC存活率，caspase-3蛋白的表达水平。结果 A、B、C组RGC存活率分别为53.63%±7.35%、57.86%±6.00%、45.00%±4.37%（F=7.11，P=0.029），其中A组与C组（P=0.025），B组和C组（P=0.002）之间均有显著性差异；A 组和B组之间无显著性差异（P=0.24）。A、B、C三组视网膜组织中Caspase-3蛋白与内参灰度比值分别为0.20±0.02、0.19±0.02、0.24±0.03（F=9.73，P=0.02）。其中A组与C组（P=0.005），B组和C组（P=0.001）之间均有显著性差异；A 组和B组之间无显著性差异（P=0.418）。结论 小干扰RNA能有效保护大鼠视神经夹伤模型的RGC，提高RGC的存活率。

Neuroprotection of siRNA interference to the retinal ganglion cells in optic nerve crush model rat

 Objective To investigate the neuroprotective effects of small interfering RNA (siRNA) on retinal ganglion cells (RGC) in optic nerve crush model rats. Design Experimental study. Participants fifty-four SD rats. Methods fifty-four SD rats were divided into 3 groups, group A, B, and C, 18 rats per group. The right eyes of all rats were crushed by micro-clipping with 40 g power for 60 s, the left eyes were not crushed as the controls. A and B group was given siRNA 10 μg/per eye and siRNA 20 μg/per eye, the group C was given normal saline intravitreal injectionon immediately after optic nerve crush. The experiment lasted for 14 days. Six rats in each group were retrograde labeled by injection of 3% fluorogold (FG) into both side of superior colliculus at 10 days after the crush. At the 14 day, the rats were sacrificed and stretched preparation of retina, RGCs were photographed and counted with mask method. The survival percentage of RGCs = RGCs density in right eye / RGCs density in left eye. Further the western blot was used to exam the expression level of caspase-3 protein in retinal tissue. Main Outcome Measures The survival percentage of RGC, the expression level of caspase-3 protein. Results The average survival percentage of RGCs in A, B and C group was 53.63%±7.35%, 57.86%±6.00%, 45.00%±4.37% respectively (F=7.11, P=0.029). A significant difference was found between group A and C(P=0.025), between group B and C(P=0.002) as well. The difference between group A and B was not significant (P=0.24). The expression level of caspase-3 protein in the A, B, C groups in the retinal tissue was 0.20±0.02, 0.19±0.02, 0.24±0.03 (F=9.73, P =0.02). A significant difference was found between group A and C(P=0.005), between group B and C(P=0.001) as well. The difference between group A and B was not significant(P=0.418). Conclusion siRNA shows the neuroprotective effects on RGCs in optic nerve crush model rats.