Experimental evidence has shown that probucol, a hypocholesterolemic agent, is also able to increase glutathione peroxidase (GPx) activity. However, there is a lack of knowledge about the mechanism(s) involved in this event. In this study, in vitro experiments with purified GPx1 from bovine erythrocytes and cultured SH-SY5Y neuroblastoma cells, as well as in silico studies with GPx1, were performed in order to elucidate mechanisms mediating the stimulatory effect of probucol on GPx activity and to investigate the relevance of this event in terms of susceptibility against peroxide-induced cytotoxicity. In vitro experiments with purified GPx1 showed a direct stimulatory effect of probucol on the activity of GPx1, which was related to an increase in V_max with no changes in K_M. Probucol also increased GPx activity in cultured SH-SY5Y neuroblastoma cells, while the levels of GPx1 expression were not changed, corroborating the results found with the purified enzyme. In addition, probucol rendered SH-SY5Y cells more resistant to hydroperoxide-induced cytotoxicity, and this event was abolished in GPx1 knocked-down cells. In silico studies with GPx1 pointed to a potential binding site for probucol at the close vicinity of the GSH pocket. Collectively, the results presented herein indicate that GPx1 plays a central role in the probucol-induced protective effects against peroxide toxicity. This highlights a novel target (GPx1) and a new mechanism of action (direct activation) for an “old drug.” The relevance of such results for in vivo conditions deserves further investigation.

实验证据表明，降胆固醇药物普罗布考也能够增加谷胱甘肽过氧化物酶 (GPx) 的活性。然而，对这一事件所涉及的机制缺乏了解。在这项研究中，进行了从牛红细胞和培养的 SH-SY5Y 神经母细胞瘤细胞中纯化的 GPx1 的体外实验，以及用 GPx1 进行的计算机研究，以阐明介导普罗布考对 GPx 活性的刺激作用的机制，并研究这一事件在对过氧化物诱导的细胞毒性的敏感性方面的相关性。用纯化的 GPx1 进行的体外实验表明，普罗布考对 GPx1 的活性有直接的刺激作用，这与V _max增加而K没有变化有关_米。普罗布考还增加了培养的 SH-SY5Y 神经母细胞瘤细胞中的 GPx 活性，而 GPx1 表达水平没有改变，证实了用纯化酶发现的结果。此外，普罗布考使 SH-SY5Y 细胞对氢过氧化物诱导的细胞毒性更具抵抗力，并且这一事件在 GPx1 敲低的细胞中被消除。用 GPx1 进行的计算机研究表明，在 GSH 口袋附近有一个潜在的普罗布考结合位点。总的来说，本文提供的结果表明 GPx1 在普罗布考诱导的针对过氧化物毒性的保护作用中起核心作用。这突出了“老药”的新靶点（GPx1）和新的作用机制（直接激活）。这些结果与体内条件的相关性值得进一步研究。