Research and Development AL.CHI.MI.A. S.r.l, Viale Austria 14, 35020, Ponte San Nicolò, PD, Italy.
Department of Biomedical Sciences, Humanitas University, Via Rita Levi Montalcini 4, 20090, Pieve Emanuele, Milano, Italy; Eye Center, Humanitas Gavazzeni-Castelli, Via Gavazzeni 21, 24125, Bergamo, Italy.
Exp Eye Res. 2021 Dec;213:108837. doi: 10.1016/j.exer.2021.108837. Epub 2021 Nov 10.
This study aimed to evaluate viability of retinal cells after the use of multiple intraoperative devices, namely a vitreal dye (triamcinolone acetonide,TA), a ERM/ILM dye (solution of trypan blue 0.15% and brilliant blue 0.025%), and two intraocular tamponades, namely perfluoro-n-octane, (PFO) and silicone oil (SO 1000 cSt), with minimal and maximal removal of their residues, during a simulated pars plana vitrectomy (PPV) in porcine eyes ex-vivo. The in vitro cytotoxicity of each of these compounds was verified on ARPE-19 cells by direct tests according to the ISO 10993-5 (2009). Pars plana vitrectomy was performed on 25 enucleated porcine eyes divided in five groups according to the following conditions: Group A) No surgery control: eye bulbs were kept at room temperature for 40 min; Group B) Sham surgery: PPV with the sole use of BSS for 40 min; Group C) Cytotoxic control: PPV with BSS infusion (20 min) followed by intravitreal injection of 1H-PFO (contact time: 20 min); Group D) Surgery with residues: PPV with BSS infusion and sequential intravitreal injection of TA, ERM/ILM dye, PFO and SO, with minimal removal of each compound after a specified contact-time (overall duration: 40 min); Group E) Surgery with minimal residues: PPV performed as in group D, but with maximal removal of each compound (overall duration: 40 min). All the experimental procedures were performed at room temperature. Immediately after surgery, the retina was extracted from each eye bulb and samples of 3-mm diameter were prepared. Retinal viability was determined for each sample by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide assay. A cell viability <70% was considered the cytotoxicity threshold. Kruskal-Wallis test was used to evaluate the differences in retinal viability between groups. No cytotoxicity was detected in retinal samples in groups A, B and E. Samples from eye bulbs that had undergone surgery with minimal removal of residues (group D) and cytotoxic controls (group C) showed high retinal cytotoxicity. The tested conditions indicated that the combined use of TA, ERM/ILM dye, PFO and SO during PPV does not affect retinal cells viability if all the devices are properly removed, whereas the cytotoxicity detected in group D may suggest that the presence and accumulation of the residues of the compounds used intraoperatively could negatively impact retinal viability due to a cumulative and/or synergistic cytotoxic effect between them, supporting the crucial role of an optimal removal of the intraoperative medical devices to ensure a safe vitrectomy to the patient.
本研究旨在评估在模拟的眼后段玻璃体切除术（PPV）中，使用多种术中器械（即玻璃体内染料[曲安奈德]、ERM/ILM 染料[曲安奈德和丽春红的混合液]和两种眼内填充物，即全氟萘烷（PFO）和硅油（SO 1000 cSt））对视网膜细胞活力的影响。每种化合物的体外细胞毒性均通过直接测试（根据 ISO 10993-5（2009））在 ARPE-19 细胞上进行了验证。对 25 只经眼窝摘除的猪眼进行 PPV，根据以下条件将其分为五组：A 组）无手术对照：眼球在室温下保持 40 分钟；B 组）假手术对照：仅用 BSS 进行 40 分钟的 PPV；C 组）细胞毒性对照：BSS 输注（20 分钟）后，玻璃体内注射 1H-PFO（接触时间：20 分钟）；D 组）有残留物的手术：BSS 输注和序贯玻璃体内注射 TA、ERM/ILM 染料、PFO 和 SO，每种化合物在指定接触时间后（总时间：40 分钟）进行最小程度的清除；E 组）最小残留物手术：如 D 组所述进行 PPV，但对每种化合物进行最大程度的清除（总时间：40 分钟）。所有实验程序均在室温下进行。手术后，立即从每个眼球中提取视网膜，并制备 3mm 直径的样本。通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐法测定每个样本的视网膜活力。细胞活力<70%被认为是细胞毒性阈值。Kruskal-Wallis 检验用于评估各组间视网膜活力的差异。A、B 和 E 组的视网膜样本未检测到细胞毒性。经过最小程度清除残留物的手术（D 组）和细胞毒性对照（C 组）的眼球样本显示出高度的视网膜细胞毒性。结果表明，在适当清除所有器械的情况下，TA、ERM/ILM 染料、PFO 和 SO 在 PPV 中联合使用不会影响视网膜细胞活力，但 D 组中检测到的细胞毒性可能表明术中使用的化合物的残留物的存在和积累可能会对视网膜活力产生负面影响，因为它们之间可能存在累积和/或协同的细胞毒性作用，这支持了为确保患者安全进行玻璃体切除术而优化清除术中医疗器械的重要作用。
