脂多糖暴露诱导秀丽隐杆线虫氧化损伤：肌肽的保护作用。
Lipopolysaccharide exposure induces oxidative damage in Caenorhabditis elegans: protective effects of carnosine.
机构信息
Department of Pharmacy, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, 200092, P. R. China.
Department of Endocrinology, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, 200092, P. R. China.
出版信息
BMC Pharmacol Toxicol. 2020 Dec 3;21(1):85. doi: 10.1186/s40360-020-00455-w.
BACKGROUND
The present study was designed to investigate the protective effects and mechanisms of carnosine on lipopolysaccharide (LPS)-induced injury in Caenorhabditis elegans.
METHODS
C. elegans individuals were stimulated for 24 h with LPS (100 μg/mL), with or without carnosine (0.1, 1, 10 mM). The survival rates and behaviors were determined. The activities of superoxide dismutase (SOD), glutathione reductase (GR), and catalase (CAT) and levels of malondialdehyde (MDA) and glutathione (GSH) were determined using the respective kits. Reverse transcription polymerase chain reaction (RT-PCR) was performed to validate the differential expression of sod-1, sod-2, sod-3, daf-16, ced-3, ced-9, sek-1, and pmk-1. Western blotting was used to determine the levels of SEK1, p38 mitogen-activated protein kinase (MAPK), cleaved caspase3, and Bcl-2. C. elegans sek-1 (km2) mutants and pmk-1 (km25) mutants were used to elucidate the role of the p38 MAPK signaling pathway.
RESULTS
Carnosine improved the survival of LPS-treated C. elegans and rescued behavioral phenotypes. It also restrained oxidative stress by decreasing MDA levels and increasing SOD, GR, CAT, and GSH levels. RT-PCR results showed that carnosine treatment of wild-type C. elegans up-regulated the mRNA expression of the antioxidant-related genes sod-1, sod-2, sod-3, and daf-16. The expression of the anti-apoptosis-related gene ced-9 and apoptosis-related gene ced-3 was reversed by carnosine. In addition, carnosine treatment significantly decreased cleaved caspase3 levels and increased Bcl-2 levels in LPS-treated C. elegans. Apoptosis in the loss-of-function strains of the p38 MAPK signaling pathway was suppressed under LPS stress; however, the apoptotic effects of LPS were blocked in the sek-1 and pmk-1 mutants. The expression levels of sek-1 and pmk-1 mRNAs were up-regulated by LPS and reversed by carnosine. Finally, the expression of p-p38MAPK and SEK1 was significantly increased by LPS, which was reversed by carnosine.
CONCLUSION
Carnosine treatment protected against LPS injury by decreasing oxidative stress and inhibiting apoptosis through the p38 MAPK pathway.
背景
本研究旨在探讨肌肽对脂多糖（LPS）诱导的秀丽隐杆线虫损伤的保护作用及其机制。
方法
用 LPS（100μg/mL）刺激秀丽隐杆线虫 24h，同时给予或不给予肌肽（0.1、1、10mM）。测定存活率和行为。采用相应试剂盒测定超氧化物歧化酶（SOD）、谷胱甘肽还原酶（GR）和过氧化氢酶（CAT）的活性以及丙二醛（MDA）和谷胱甘肽（GSH）的水平。采用反转录聚合酶链反应（RT-PCR）验证 sod-1、sod-2、sod-3、daf-16、ced-3、ced-9、sek-1 和 pmk-1 的差异表达。采用 Western 印迹法测定 SEK1、p38 丝裂原活化蛋白激酶（MAPK）、cleaved caspase3 和 Bcl-2 的水平。采用 sek-1（km2）突变体和 pmk-1（km25）突变体阐明 p38 MAPK 信号通路的作用。
结果
肌肽提高 LPS 处理的秀丽隐杆线虫的存活率并挽救行为表型。它还通过降低 MDA 水平和增加 SOD、GR、CAT 和 GSH 水平来抑制氧化应激。RT-PCR 结果表明，肌肽处理野生型秀丽隐杆线虫可上调抗氧化相关基因 sod-1、sod-2、sod-3 和 daf-16 的 mRNA 表达。肌肽逆转了凋亡相关基因 ced-3 和 ced-9 的表达。此外，肌肽处理可显著降低 LPS 处理的秀丽隐杆线虫中 cleaved caspase3 的水平并增加 Bcl-2 的水平。在 LPS 应激下，p38 MAPK 信号通路的功能丧失株系中的细胞凋亡受到抑制；然而，LPS 的凋亡作用在 sek-1 和 pmk-1 突变体中被阻断。LPS 上调 sek-1 和 pmk-1 mRNA 的表达水平，肌肽逆转了这一作用。最后，LPS 显著增加了 p-p38MAPK 和 SEK1 的表达，肌肽逆转了这一作用。
结论
肌肽通过降低氧化应激和抑制凋亡来保护秀丽隐杆线虫免受 LPS 损伤，其作用机制与 p38 MAPK 途径有关。
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