Nishimoto Shoichi, Koike Shin, Inoue Naho, Suzuki Toshihiro, Ogasawara Yuki
Department of Analytical Biochemistry, Meiji Pharmaceutical University, 2-522-1 Noshio, Kiyose, Tokyo, 204-8588, Japan.
Department of Analytical Biochemistry, Meiji Pharmaceutical University, 2-522-1 Noshio, Kiyose, Tokyo, 204-8588, Japan.
Biochem Biophys Res Commun. 2017 Feb 5;483(2):874-879. doi: 10.1016/j.bbrc.2017.01.024. Epub 2017 Jan 7.
The present study focused on the methylglyoxal (MG) detoxification mechanism in neuroblastoma cells. The involvement of nuclear factor erythroid 2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) pathway as a defense response against the formation of MG-modified proteins, which is well-known evidence of carbonyl stress, was also examined. We found that MG treatment resulted in accumulation of modified proteins bearing the structure of advanced glycation end products (AGEs) derived from MG in SH-SY5Y cells. This accumulation was suppressed by activation of the Nrf2 pathway prior to MG exposure via pre-treatment with an Nrf2 activator, carnosic acid and CDDO-Im, confirming the involvement of the Nrf2 pathway in MG detoxification. Although pre-treatment with the Nrf2 activator did not affect mRNA levels of GLO1, AKR1B1, and AKR7A2, the expressions of GCL and xCT mRNA, involved in GSH synthesis, were induced prior to increase in GSH levels. Furthermore, we demonstrated that a GSH synthesis inhibitor eliminated the MG detoxification effect derived from pretreatment with the Nrf2 activator. These results indicated that increase in GSH levels, induced by pre-treatment with carnosic acid, promoted the formation of the GLO1 substrate, hemithioacetal, thereby accelerating MG metabolism via the glyoxalase system and suppressing its toxicity. It was, therefore, determined that promotion of GSH synthesis via the Nrf2/Keap1pathway is important in the MG detoxification mechanism against neuronal MG-induced carbonyl stress, and Nrf2 activators contribute to reduction in the accumulation and toxic expression of carbonyl proteins.
本研究聚焦于神经母细胞瘤细胞中的甲基乙二醛（MG）解毒机制。还研究了核因子红细胞2相关因子2（Nrf2）/ Kelch样ECH相关蛋白1（Keap1）途径作为针对MG修饰蛋白形成的防御反应的参与情况，这是羰基应激的著名证据。我们发现，MG处理导致SH-SY5Y细胞中积累了具有源自MG的晚期糖基化终产物（AGEs）结构的修饰蛋白。通过用Nrf2激活剂肌醇六磷酸和CDDO-Im进行预处理，在MG暴露之前激活Nrf2途径可抑制这种积累，证实了Nrf2途径参与MG解毒。尽管用Nrf2激活剂进行预处理不影响GLO1、AKR1B1和AKR7A2的mRNA水平，但参与谷胱甘肽（GSH）合成的GCL和xCT mRNA的表达在GSH水平升高之前就被诱导了。此外，我们证明GSH合成抑制剂消除了源自Nrf2激活剂预处理的MG解毒作用。这些结果表明，肌醇六磷酸预处理诱导的GSH水平升高促进了GLO1底物半硫代缩醛的形成，从而通过乙二醛酶系统加速MG代谢并抑制其毒性。因此，确定通过Nrf2 / Keap1途径促进GSH合成在针对神经元MG诱导的羰基应激的MG解毒机制中很重要，并且Nrf2激活剂有助于减少羰基蛋白的积累和毒性表达。
