Kim In-Hye, Choi Jeong-Wook, Nam Taek-Jeong
Cell Biology Laboratory, Institute of Fisheries Sciences, Pukyong National University, Busan 46041, Republic of Korea.
Department of Food Science and Nutrition, Pukyong National University, Busan 48513, Republic of Korea.
Exp Ther Med. 2020 Feb;19(2):849-860. doi: 10.3892/etm.2019.8304. Epub 2019 Dec 9.
Acetaminophen (APAP) is a widely used analgesic and antipyretic. It is safe at normal treatment doses; however, APAP overdose is a major cause of acute liver and kidney failure. A variety of methods to reduce the damage caused by APAP overdose have previously been evaluated. The protein-rich seaweed has antioxidant, antitumor and anti-inflammatory activities, and protects against cytotoxicity. However, little is known regarding the protective effects of peptide against APAP-induced hepatotoxicity. The present study investigated the ability of peptide (PYP1-4) to ameliorate the damage caused by APAP-induced hepatotoxicity using HepG2 as the model cell line in addition to the signaling pathways involved. Briefly, cell viability, nitric oxide, reactive oxygen species and apoptosis assays were performed in conjunction with western blot analysis and reverse transcription-quantitative PCR. First, the present study revealed the minimum toxic concentration of APAP (15 mM) and the resting concentration of PYP1-4 (0-500 ng/ml). Administration of PYP1-4 to APAP-induced cells decreased the nitric oxide and reactive oxygen species levels, and restored the levels of antioxidant-associated proteins (catalase, heme oxygenase 1, superoxide dismutase 2 and quinone oxidoreductase 1). PYP1-4 increased the translocation of nuclear factor, erythroid 2 like 2 to the nucleus and the activities of glycogen synthase kinase-3β, Akt and AMP-activated protein kinase. In addition, APAP induced apoptosis; however, PYP1-4 inhibited apoptosis by modulating the levels of pro-apoptotic markers (Bad), anti-apoptotic markers (Bcl-2 and BH3 interacting domain death agonist), caspases and poly (ADP-ribose) polymerase 1. Subsequently, the insulin-like growth factor 1 receptor signaling pathway was investigated to determine whether PYP1-4 treatment restored the levels of cell growth-associated factors during APAP-induced hepatotoxicity. PYP1-4 treatment impacted the levels of components of the insulin receptor substrate 1/PI3K/Akt and Ras/Raf/ERK signaling pathways, and promoted cell survival. Therefore, the peptide PYP1-4 may be useful for preventing APAP-induced hepatotoxicity.
对乙酰氨基酚（APAP）是一种广泛使用的镇痛和解热药。在正常治疗剂量下它是安全的；然而，APAP过量是急性肝衰竭和肾衰竭的主要原因。此前已经评估了多种减少APAP过量所致损伤的方法。富含蛋白质的海藻具有抗氧化、抗肿瘤和抗炎活性，并能抵御细胞毒性。然而，关于肽对APAP诱导的肝毒性的保护作用知之甚少。本研究以HepG2作为模型细胞系，研究了肽（PYP1-4）改善APAP诱导的肝毒性所造成损伤的能力以及相关信号通路。简而言之，结合蛋白质印迹分析和逆转录-定量PCR进行细胞活力、一氧化氮、活性氧和凋亡检测。首先，本研究揭示了APAP的最小毒性浓度（15 mM）和PYP1-4的静息浓度（0-500 ng/ml）。将PYP1-4给予APAP诱导的细胞可降低一氧化氮和活性氧水平，并恢复抗氧化相关蛋白（过氧化氢酶、血红素加氧酶1、超氧化物歧化酶2和醌氧化还原酶1）的水平。PYP1-4增加了核因子红细胞2样2向细胞核的转位以及糖原合酶激酶-3β、Akt和AMP活化蛋白激酶的活性。此外，APAP诱导细胞凋亡；然而，PYP1-4通过调节促凋亡标志物（Bad）、抗凋亡标志物（Bcl-2和BH3相互作用结构域死亡激动剂）、半胱天冬酶和聚（ADP-核糖）聚合酶1的水平来抑制细胞凋亡。随后，研究了胰岛素样生长因子1受体信号通路，以确定PYP1-4处理是否能在APAP诱导的肝毒性期间恢复细胞生长相关因子的水平。PYP1-4处理影响胰岛素受体底物1/PI3K/Akt和Ras/Raf/ERK信号通路成分的水平，并促进细胞存活。因此，肽PYP1-4可能有助于预防APAP诱导的肝毒性。
