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目的 利用玻璃体腔注射β淀粉样蛋白(amyloid β,Aβ)制作视网膜炎症模型.方法 实验研究.分别向25只C57BL/6J小鼠的玻璃体腔注射2μl PBS或不同浓度(0.5、1.0、1.5、2.0 μg/μl)的Aβ1-40 2μl,24 h后检测视网膜神经层(简称为神经视网膜)中白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)和NLRP3炎性小体的mRNA水平变化确定适宜的Aβ诱导浓度.确定浓度后,分别向小鼠玻璃体腔注射2 μl最适浓度的Aβ40-1 (n=12)、Aβ1-40(n=12),于第1、4、14天检测神经视网膜和视网膜色素上皮(RPE)/脉络膜复合体中IL-6、TNF-α和NLRP3的mRNA水平变化;分别向小鼠玻璃体腔注射2 μl最适浓度的PBS(n=3)、Aβ40-1 (n=3)和Aβ1-40(n=3),于第4天取眼球行HE染色,光镜下观察视网膜形态改变.于第4、14天对正常对照组(n=12)、Aβ40-1组(n=8)和Aβ1-40组(n=8)行全视野视网膜电图(ERG)检测视网膜功能.用方差分析对各参数进行统计学处理.结果 诱导视网膜炎症损伤模型的最适Aβ1-40浓度为1.0μg/μl.与注射Aβ40-1组相比,Aβ1-40组在神经视网膜和RPE/脉络膜复合体中的IL-6、TNF-α和NLRP3炎性小体的mRNA水平在第1、4天均有明显升高且差异具有统计学意义.而在第14天时,除神经视网膜中的TNF-α外,神经视网膜和RPE/脉络膜复合体中的各炎性因子mRNA水平的差异均无统计学意义.第4天时,各组HE染色均未发现视网膜有明显的结构紊乱和炎性细胞浸润.玻璃体腔注射后第4、14天时,Aβ1-40组小鼠的ERG a波和b波振幅明显低于正常对照组和Aβ40-1组且差异具有明显统计学意义;而第14天时正常对照组和Aβ40-1组小鼠的ERG振幅差别除在暗适应光刺激强度为1.0、10.0 cd·s/m2以外,在其余条件下差异均无统计学意义.结论 玻璃体腔注射Aβ1-40可诱导神经视网膜和RPE/脉络膜复合体产生一过性炎症反应并使视网膜产生功能障碍,但视网膜结构未出现明显变化.炎性小体在这一过程中被活化,这些特征符合早期年龄相关性黄斑变性(AMD)的主要病理特征,该模型可用于早期AMD的相关研究.

Objective To establish a mouse model of retinal inflammation induced by the intravitreal injection of amyloid β (Aβ), a major component in drusen.Methods This was an experimental study in C57BL/6J mice to determine the optimal concentration of Aβ1-40, the mRNA expressions of inflammatory cytokines interleukin-6 (IL-6), the tumor necrosis factor-α (TNF-α) and nucleotide-binding oligomerization domain leucine-rich repeats containing pyrin domain 3 (NLRP3) inflammasome were examined at 24 hours after the intravitreal injection of 2 μl of PBS or different concentrations (0.5, 1.0, 1.5, 2.0 μg/μl) of Aβ1-40.After determination of the concentration, mice were injected with optimal concentrations of Aβ1-40 (n=12) or Aβ40-1 (n=12) and the eyeballs were enucleated at days 1, 4 and 14 postinjection.The expression of inflammatory cytokines and the activation of NLRP3 inflammasome in the neuroretina and the RPE/choroid complex were analyzed by real-time PCR.The severity of retinal damage was evaluated with HE staining at day 4 postinjection (n=3 for each group), and retinal functions were evaluated by ERG at days 4 and 14 (n=12 in PBS group, n=8 in Aβ40-1 group and Aβ1-40 group).Differences between groups were analyzed using ANOVA followed by a post hoc Bonferroni correction.Results The optimal concentration of Aβ1-40 that could induce retinal inflammation was 1.0 μg/μl.Compared with Aβ40-1, Aβ1-40 significantly upregulated the expressions of IL-6, TNF-α and NLRP3 inflammasome genes in the neuroretina and the RPE/choroid complex at days 1 and 4 postinjection.The differences in inflammatory cytokine gene expression between these groups were not significant at day 14, except for the TNF-α in the neuroretina group.No structural disorganization or inflammatory cell infiltration was observed in the retina in each group under light microscopy at day 4.At days 4 and 14 after intravitreal injection, the amplitudes of ERG a-and b-waves significantly decreased in the Aβ1-40 injected mice compared to the untreated and the Aβ40-1 injected mice.No significant differences were observed between the untreated and Aβ40-1 groups at day 14, except for the amplitudes of the dark-adapted ERG b-wave recorded with light intensities of 1.0 and 10.0 cd·s/m2.Conclusion Our data show that Aβ1-40 induces a transient proinflammatory response in the neuroretina and the RPE/choroid complex, and causes retinal functional impairment.These responses mimic those observed in the early stages of age-related macular degeneration (AMD) patients.We find inflammasome is activated in this process.This novel model is useful for investigating the pathogeneses of early AMD.