Noack Volker, Pak Kwang, Jalota Rahul, Kurabi Arwa, Ryan Allen F
Department of Surgery and Otolaryngology, School of Medicine, University of California, San Diego, La JollaCA, United States.
VA San Diego Healthcare System, San DiegoCA, United States.
Front Cell Neurosci. 2017 Aug 18;11:242. doi: 10.3389/fncel.2017.00242. eCollection 2017.
Reactive oxygen species are important elements in ototoxic damage to hair cells (HCs), appearing early in the damage process. Higher levels of natural antioxidants are positively correlated with resistance to ototoxins and many studies have shown that exogenous antioxidants can protect HCs from damage. While a very wide variety of antioxidants with different characteristics and intracellular targets exist, most ototoxicity studies have focused upon one or a few well-characterized compounds. Relatively little research has attempted to determine the comparative efficacy of large variety of different antioxidants. This has been in part due to the lack of translation between cell culture and measures of efficacy. To circumvent this limitation, we used an assay based on micro-explants from the basal and middle turns of the neonatal mouse organ of Corti to screen a commercial redox library of diverse antioxidant compounds for their ability to protect mammalian HCs from a high dose of the ototoxic antibiotic gentamicin. The library included several antioxidants that have previously been studied as potential treatments for HC damage, as well as many antioxidants that have never been applied to ototoxicity. The micro-explants were treated with 200 μM gentamicin alone, gentamicin plus one of three dosages of a redox compound, the highest dosage of compound alone, or were untreated. HC counts were determined before the gentamicin insult and at 1, 2, and 3 days afterward to evaluate the HC survival. From a total of 81 antioxidant compounds, 13 exhibited significant protection of HCs. These included members of a variety of antioxidant classes with several novel antioxidants, not previously tested on HCs, appearing to alleviate the damaging gentamicin effect. Some compounds previously shown to be protective of HCs were correspondingly protective in this screen, while others were not. Finally, one of the three pro-oxidant compounds included in the library as well as six antioxidants exhibited evidence of toxicity in the absence of gentamicin. The results demonstrate the wide variability in the ability of antioxidants to protect HCs from high-dose gentamicin damage, and identify promising candidate leads for further study as potential drug targets.
• A medium-throughput assay based on micro-explants of the organ of Corti was developed to screen mammalian cochlear hair cells for protection from damage by ototoxins.• Eighty one antioxidants and 3 pro-oxidants were evaluated for hair cell protection from high-dose gentamicin.• Thirteen antioxidants were significantly protective, while 6 proved to be damaging.• The use of a common assay permitted an evaluation of the relative capacity of different antioxidants for the protection of hair cells.
活性氧是毛细胞耳毒性损伤的重要因素，在损伤过程中早期出现。较高水平的天然抗氧化剂与对耳毒素的抗性呈正相关，许多研究表明外源性抗氧化剂可以保护毛细胞免受损伤。虽然存在各种各样具有不同特性和细胞内靶点的抗氧化剂，但大多数耳毒性研究都集中在一种或几种特征明确的化合物上。相对较少的研究试图确定多种不同抗氧化剂的比较功效。部分原因是细胞培养与功效测量之间缺乏转化。为了克服这一限制，我们使用了一种基于新生小鼠柯蒂氏器基部和中部微切片的试验，来筛选一个包含多种抗氧化化合物的商业氧化还原文库，以评估它们保护哺乳动物毛细胞免受高剂量耳毒性抗生素庆大霉素损伤的能力。该文库包括几种先前作为毛细胞损伤潜在治疗方法进行过研究的抗氧化剂，以及许多从未应用于耳毒性研究的抗氧化剂。微切片分别用单独的200μM庆大霉素、庆大霉素加三种剂量之一的氧化还原化合物、单独的最高剂量化合物处理，或不进行处理。在庆大霉素损伤前以及损伤后1、2和3天确定毛细胞数量，以评估毛细胞存活情况。在总共81种抗氧化化合物中，有13种对毛细胞有显著保护作用。这些包括各种抗氧化剂类别中的成员，有几种以前未在毛细胞上测试过的新型抗氧化剂，似乎减轻了庆大霉素的损伤作用。一些先前显示对毛细胞有保护作用的化合物在本次筛选中相应地具有保护作用，而其他化合物则没有。最后，文库中包含的三种促氧化化合物之一以及六种抗氧化剂在没有庆大霉素的情况下表现出毒性证据。结果表明抗氧化剂保护毛细胞免受高剂量庆大霉素损伤的能力差异很大，并确定了有前景的候选先导物，作为进一步研究的潜在药物靶点。
• 开发了一种基于柯蒂氏器微切片的中通量试验，以筛选哺乳动物耳蜗毛细胞免受耳毒素损伤的保护作用。
• 评估了81种抗氧化剂和3种促氧化剂对毛细胞免受高剂量庆大霉素损伤的保护作用。
• 13种抗氧化剂具有显著保护作用，而6种被证明具有损伤作用。
• 使用共同的试验允许评估不同抗氧化剂保护毛细胞的相对能力。
