Division of Pharmacology and Pharmacotherapy, Faculty of Pharmacy, University of Helsinki, Finland.
Division of Pharmacology and Pharmacotherapy, Faculty of Pharmacy, University of Helsinki, Finland; Integrative Physiology and Pharmacology Unit, Institute of Biotechnology, Faculty of Medicine, University of Turku, Finland; School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Finland.
Free Radic Biol Med. 2021 Jun;169:14-23. doi: 10.1016/j.freeradbiomed.2021.04.001. Epub 2021 Apr 7.
Oxidative stress (OS) is a common toxic feature in various neurodegenerative diseases. Therefore, reducing OS could provide a potential approach to achieve neuroprotection. Prolyl oligopeptidase (PREP) is a serine protease that is linked to neurodegeneration, as endogenous PREP inhibits autophagy and induces the accumulation of detrimental protein aggregates. As such, inhibition of PREP by a small-molecular inhibitor has provided neuroprotection in preclinical models of neurodegenerative diseases. In addition, PREP inhibition has been shown to reduce production of reactive oxygen species (ROS) and the absence of PREP blocks stress-induced ROS production. However, the mechanism behind PREP-related ROS regulation is not known. As we recently discovered PREP's physiological role as a protein phosphatase 2A (PP2A) regulator, we wanted to characterize PREP inhibition as an approach to reduce OS. We studied the impact of a PREP inhibitor, KYP-2047, on hydrogen peroxide and ferrous chloride induced ROS production and on cellular antioxidant response in HEK-293 and SH-SY5Y cells. In addition, we used HEK-293 and SH-SY5Y PREP knock-out cells to validate the role of PREP on stress-induced ROS production. We were able to show that absence of PREP almost entirely blocks the stress-induced ROS production in both cell lines. Reduced ROS production and smaller antioxidant response was also seen in both cell lines after PREP inhibition by 10 μM KYP-2047. Our results also revealed that the OS reducing mechanism of PREP inhibition is related to reduced activation of ROS producing NADPH oxidase through enhanced PP2A activation. In conclusion, our results suggest that PREP inhibition could also provide neuroprotection by reducing OS, thus broadening the scope of its beneficial effects on neurodegeneration.
氧化应激（OS）是各种神经退行性疾病的共同毒性特征。因此，减少 OS 可能是实现神经保护的一种潜在方法。脯氨酰寡肽酶（PREP）是一种与神经退行性变有关的丝氨酸蛋白酶，内源性 PREP 抑制自噬并诱导有害蛋白聚集体的积累。因此，小分子抑制剂抑制 PREP 在神经退行性疾病的临床前模型中提供了神经保护作用。此外，已经表明 PREP 抑制可减少活性氧物种（ROS）的产生，并且 PREP 的缺失阻止了应激诱导的 ROS 产生。但是，与 PREP 相关的 ROS 调节的机制尚不清楚。由于我们最近发现 PREP 作为蛋白磷酸酶 2A（PP2A）调节剂的生理作用，因此我们希望将 PREP 抑制作为减少 OS 的一种方法进行研究。我们研究了 PREP 抑制剂 KYP-2047 对过氧化氢和氯化亚铁诱导的 ROS 产生以及对 HEK-293 和 SH-SY5Y 细胞中细胞抗氧化反应的影响。此外，我们使用 HEK-293 和 SH-SY5Y PREP 敲除细胞来验证 PREP 在应激诱导的 ROS 产生中的作用。我们能够表明，在两种细胞系中，PREP 的缺失几乎完全阻止了应激诱导的 ROS 产生。在用 10μM KYP-2047 抑制 PREP 后，两种细胞系中的 ROS 产生减少和抗氧化反应也较小。我们的结果还表明，PREP 抑制的 OS 减少机制与通过增强 PP2A 激活来减少 ROS 产生的 NADPH 氧化酶的激活有关。总之，我们的结果表明，通过减少 OS，PREP 抑制还可以提供神经保护作用，从而扩大了其对神经退行性变有益作用的范围。
