The Effect of Preventing Oxidative Stress and Its Mechanisms in the Extract from Sonchus brachyotus DC. Based on the Nrf2-Keap1-ARE Signaling Pathway

Abstract

As the organ with the largest contact area with the outside world, the intestine is home to a large number of microorganisms and carries out the main functions of food digestion, absorption, and metabolism. Therefore, there is a very active metabolism of substances and energy in the gut, which is easily attacked by oxygen free radicals. What is more, oxidative stress can gradually and slowly cause very serious damage to the gut. Hence, maintaining redox balance is essential for maintaining environmental balance in the gut. Our previous studies have demonstrated that the extract of Sonchus brachyotus DC. (SBE) has been shown to be capable of repairing oxidative damage, while it has not been demonstrated that it can prevent oxidative stress or how it develops. In this work, we investigated the prevention of oxidative stress and its mechanism in SBE based on the H₂O₂-induced oxidative damage model in Caco-2 cells; the results indicate that SBE can reduce the contents of ROS and MDA and increase the activities of SOD and CAT in preventing oxidative stress. Then, at the mRNA and protein level, SBE can up-regulate and down-regulate the expression of related genes (NFE2L2, KEAP1, HMOX1, NQO1, SOD1, CAT, and GPX1) and proteins involved in the Nrf2-Keap1-ARE signaling pathway. In conclusion, SBE plays a preventive role in oxidative stress through the Nrf2-Keap1-ARE signaling pathway.

Keywords: Nrf2-Keap1-ARE signaling pathway; Sonchus brachyotus DC. extracts (SBE); preventing oxidative stress.

Figure 1

Effect of SBE on intracellular ROS production, MDA content, and SOD and CAT activity. (A) Cell viability; (B) ROS content; (C) MDA content; (D) CAT activity; (E) SOD activity. The values are expressed as the mean ± SD (n ≥ 3 per group). Significance between groups was analyzed using one-way ANOVA (compared with the control group, # p < 0.05, ## p < 0.01; compared with the model group, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Figure 2

Time–effect relationship of preventing oxidative stress effect of SBE. (A) ROS content; (B) MDA content; (C) CAT activity; (D) SOD activity. The values are expressed as the mean ± SD (n ≥ 3 per group). Significance between groups was analyzed using one-way ANOVA (compared with the control group, ^# p < 0.05, ^## p < 0.01; compared with the model group, * p < 0.05, ** p < 0.01).

Figure 3

Effect of SBE on Nrf2 and Keap1 mRNA and protein expression levels in cells. GAPDH was used as an internal control. (A) Expression of NFE2L2 gene; (B) Expression of KEAP1 gene; (C) Western Blot analysis of Nrf2 and keap1; (D) Expression of Nrf2 protein; (E) Expression of Keap1 protein. The values are expressed as the mean ± SD (n ≥ 3 per group). Significance between groups was analyzed using one-way ANOVA (compared with the control group, ^# p < 0.05, ^## p < 0.01; compared with the model group, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Figure 4

Effects of SBE on genes downstream of the antioxidant pathway. (A) Expression of HMOX1 gene; (B) Expression of NQO1 gene; (C) Expression of SOD1 gene; (D) Expression of CAT gene; (E) Expression of GPX1 gene; The values are expressed as the mean ± SD (n ≥ 3 per group). Significance between groups was analyzed using one-way ANOVA (compared with the control group, ^## p < 0.01; compared with the model group, * p < 0.05, ** p < 0.01, **** p < 0.0001).

Figure 5

Nrf2-Keap1-ARE signaling pathway.