Kearns S, Dawson R
Department of Pharmacodynamics, College of Pharmacy, University of Florida, Gainesville, USA.
Adv Exp Med Biol. 2000;483:563-70. doi: 10.1007/0-306-46838-7_60.
Taurine has been shown to be an effective scavenger of hypochlorous acid (HOCl). The role of HOCl is well established in tissue damage associated with reperfusion injury mediated by neutrophils. The role of HOCl in CNS injury and inflammatory reactions has not been well established. Myeloperoxidase activity is present in the CNS and it has been associated with ischemic injury. The aim of the present study was to determine the cytotoxicity of HOCl in a neuronal cell line (PC12) and the ability of taurine to prevent or reverse neurotoxicity. PC12 cells were grown in 96 well plates at a plating density of approximately 100,000 cells per well. HOCl was made up fresh from NaOCl for each experiment and the concentration verified spectrophotometrically. PC12 cells were exposed to HOCl for 1 hour in phosphate-buffered saline. Taurine was added at the time of HOCl treatment and in some experiments a post-treatment with taurine was performed by adding 1 or 10 mM taurine to the culture media (RPMI 1640). The cells were allowed 24 hours to recover and viability was determined using a tetrazolium-based (MTT) assay. The first series of experiments evaluated the toxicity of HOCl and the efficacy of taurine to protect PC12 cells. HOCl at 50 microM reduced PC12 cell viability by 50% and 150 microM reduced viability to <25% of control levels. Taurine (0.5-20 mM) was tested for cytoprotection against 150 microM HOCl and PC12 cells treated with 0.5 mM taurine exhibited only a 20% reduction in viability compared to untreated controls. Taurine concentrations of 1 mM or higher provided nearly 100% protection against HOCl. A second study was performed comparing taurine to beta-alanine, glutathione and isethionic acid. HOCl (100 microM) reduced viability to 25 +/- 1% of controls and taurine, beta-alanine and glutathione at 1 mM provided nearly complete protection. In contrast, isethionic acid, which lacks an amino group, failed to provide protection. Taurine (1 or 10 mM) added after 50 microM HOCl treatment did not provide any protection and PC12 cell viability was reduced to <39% of controls. In contrast, if taurine (50 microM) was present during the HOCl treatment and 1 mM taurine was added after the treatment, PC12 cell viability was 80 +/- 5% of controls. A combination of 250 microM taurine during the HOCl treatment and 1 mM taurine post-treatment produced 100% protection. These results clearly show that taurine is an efficient scavenger of HOCl and can prevent neuronal damage caused by HOCl. Since myeloperoxidase expression in the CNS is increased by ischemia, one function of taurine released during an ischemic event may be to scavenge HOCl and provide neuroprotection.
牛磺酸已被证明是一种有效的次氯酸（HOCl）清除剂。HOCl在由中性粒细胞介导的再灌注损伤相关的组织损伤中的作用已得到充分证实。HOCl在中枢神经系统损伤和炎症反应中的作用尚未完全明确。髓过氧化物酶活性存在于中枢神经系统中，并且与缺血性损伤有关。本研究的目的是确定HOCl对神经元细胞系（PC12）的细胞毒性以及牛磺酸预防或逆转神经毒性的能力。PC12细胞以每孔约100,000个细胞的接种密度接种于96孔板中。每次实验时用次氯酸钠新鲜配制HOCl，并通过分光光度法验证其浓度。PC12细胞在磷酸盐缓冲盐水中暴露于HOCl 1小时。在HOCl处理时加入牛磺酸，在一些实验中，通过向培养基（RPMI 1640）中加入1或10 mM牛磺酸进行牛磺酸后处理。让细胞恢复24小时，并使用基于四唑盐的（MTT）测定法测定细胞活力。第一系列实验评估了HOCl的毒性以及牛磺酸保护PC12细胞的功效。50 microM的HOCl使PC12细胞活力降低50%，150 microM使活力降低至对照水平的<25%。测试了牛磺酸（0.5 - 20 mM）对150 microM HOCl的细胞保护作用，与未处理的对照相比，用0.5 mM牛磺酸处理的PC12细胞活力仅降低了20%。1 mM或更高浓度的牛磺酸提供了几乎100%的HOCl防护。进行了第二项研究，将牛磺酸与β-丙氨酸、谷胱甘肽和羟乙磺酸进行比较。100 microM的HOCl使活力降低至对照的25 +/- 1%，1 mM的牛磺酸、β-丙氨酸和谷胱甘肽提供了几乎完全的保护。相比之下，缺乏氨基的羟乙磺酸未能提供保护。在50 microM HOCl处理后加入牛磺酸（1或10 mM）未提供任何保护，PC12细胞活力降低至对照的<39%。相反，如果在HOCl处理期间存在牛磺酸（50 microM）并在处理后加入1 mM牛磺酸，PC12细胞活力为对照的80 +/- 5%。在HOCl处理期间加入250 microM牛磺酸并在处理后加入1 mM牛磺酸可产生100%的保护。这些结果清楚地表明，牛磺酸是HOCl的有效清除剂，可以预防由HOCl引起的神经元损伤。由于缺血会增加中枢神经系统中髓过氧化物酶的表达，缺血事件期间释放的牛磺酸的一个功能可能是清除HOCl并提供神经保护。
