Safety evaluation of subretinal injection of trypan blue in rats
Safety evaluation of subretinal injection of trypan blue in rats
复制标题
大鼠视网膜下注射台盼蓝的安全性评价
DOI：
10.26355/eurrev_201805_15046
复制
发表时间：
2018-05-01
影响因子：
3.3
通讯作者：
Sun, X-H.
中科院分区：
文献类型：
--
作者：
Fang, Y.;Yao, X-Q.;Sun, X-H.
目的：确定大鼠视网膜下注射台盼蓝(TB)的适宜浓度，并提供一种在保持染料可见性的同时限制视网膜毒性的安全方案。材料和方法：成年大鼠视网膜下注射不同浓度的TB或磷酸盐缓冲盐水(PBS)，另设假注射组作为对照。在手术显微镜下观察注射区域。用视网膜电流图(ERG)检测视网膜功能。然后处死动物，对眼睛进行切片和光学显微镜检查。结果：视网膜下注射1天后，0.2%、0.08%和0.04%TB组在手术显微镜下可见TB染色，而0.02%TB组未见TB染色。注射0.2%、0.08%和0.04%Tb 2周以上组大鼠视网膜和巩膜均可见Tb染色。然而，0.2%TB注射组Erga波和b波的波幅受到影响，明显低于PBS组或假注射组。注射0.2%和0.08%Tb组大鼠视网膜外核层(ONL)、神经节细胞层(GCL)和视网膜色素上皮层(RPE)在注射后1d和7d均可见TUNEL+细胞。相比之下，0.04%Tb或PBS注射组仅有极少量的TUNEL+细胞。注射后2周，0.2%Tb注射组ONL较0.04%Tb、PBS或假注射组明显变薄。结论：Tb注射对视网膜细胞具有剂量依赖性的神经毒性作用。视网膜下注射0.04%TB是相对安全有效的视网膜下染色方法。
OBJECTIVE: To determine the appropriate concentration of trypan blue (TB) for subretinal injection in a rat model and to provide a safety profile that limits retinal toxicity while maintaining dye visibility.MATERIALS AND METHODS: Adult rats were subretinally injected with various concentrations of either TB or phosphate-buffered saline (PBS); rats which received sham injections served as an additional control. The injected areas were visualized under a surgical microscope. Electroretinography (ERG) was performed to measure retinal function. Animals were then sacrificed, and the eyes were sectioned and examined by light microscopy. Terminal deoxynucleotidy1 transferase dUTP nick-end labeling (TUNEL) was applied to determine retinal apoptosis.RESULTS: One day after the subretinal injection, TB stains were visible under the surgical microscope in the 0.2%, 0.08%, and 0.04% TB-injected groups, but not in the 0.02% TB-injected group. TB stain was detectable in the retina and sclera of the 0.2%, 0.08%, and 0.04% TB-injected groups for over 2 weeks after injection. However, the amplitudes of ERGa- and b-waves were affected and became significantly lower in the 0.2% TB-injected group than the amplitudes in the PBS-, or sham-injected group. Moreover, TUNEL+ cells appeared in the outer nuclear layer (ONL), ganglion cell layer (GCL), and retinal pigment epithelium (RPE) layer of the 0.2% and 0.08% TB-injected groups at 1 and 7 days after subretinal injection. In contrast, very few TUNEL+ cells were found in the 0.04% TB-or PBS-injected group. Two weeks after injection, the ONL was significantly thinner in the 0.2% TB-injected group than in the 0.04% TB-, PBS- or sham-injected group.CONCLUSIONS: TB injection induces a dose-dependent neurotoxic effect on retinal cells. Subretinal injection of 0.04% TB is relatively safe and effective for subretinal staining.