Abstract
Alfalfa polysaccharide (APS) is a bioactive component extracted from alfalfa that exhibits potent antioxidant properties. However, the cellular and molecular mechanisms underlying these properties remain unclear. To explore the molecular mechanism by which APS exerts antioxidant effects, an H2O2-induced oxidative stress mouse embryonic fibroblast (MEF) model was established. Cell proliferation, antioxidant enzyme activity, immune cytokine expression, and related protein expression were examined in APS-supplemented or non-supplemented conditions. The results suggested that APS strengthened the antioxidative capacity of MEFs, increasing cell proliferation, superoxide dismutase activity (SOD), and the total antioxidant capacity (T-AOC). In addition, APS reduced the secretion of interleukin (IL)-6 and IL-8 as well as expression of the proinflammatory gene retinoic acid-inducible gene I (RIG-I). APS was also able to activate the mitogen-activated protein kinase (MAPK) pathway, which promoted the translocation of nuclear factor erythroid 2–related factor 2 (Nrf2) to the nucleus. However, expression of nuclear factor-κB (NF-κB) was decreased after APS treatment. Overall, these results suggest that APS relieves H2O2-induced oxidative stress in MEFs by activating MAPK/Nrf2 signaling and suppressing NF-κB signaling. To the best of our knowledge, this is the first study to link APS with MAPK/Nrf2, NF-κB and RIG-I, thus providing new perspectives regarding the mechanisms of the antioxidant activity of APS.
Highlights
Oxidative stress, which results from an imbalance between oxidants and reductants at the cellular or organismal level, plays an important role in the development of numerous types of diseases, such as pneumonia, enteritis, and sepsis[1]
Our results strongly suggested that the proteins Toll-like receptor 4 (TLR4), myeloid differentiation primary response 88 (MyD88), IRAK1 and TRAF6 were upregulated by Alfalfa polysaccharide (APS)
These results suggest that APS can protect cells against H2O2-induced oxidative stress by activating the mitogen-activated protein kinase (MAPK)/Nrf[2] signaling pathway, which is in line with previous reports[40,41]
Summary
Oxidative stress, which results from an imbalance between oxidants and reductants at the cellular or organismal level, plays an important role in the development of numerous types of diseases, such as pneumonia, enteritis, and sepsis[1]. Most cells have evolved intricate mechanisms to prevent the generation of reactive oxygen species (ROS) or to detoxify ROS via the activation of antioxidant/detoxification enzymes, which enhance cellular ROS scavenging capacity to maintain cellular redox homeostasis and reduce oxidative damage; the balance between ROS production and antioxidant defense determines the degree of oxidative stress. The most widely studied type of oxidative stress involving protein modification is the formation of carbonyl derivatives[2,3]. Previous studies have found that alfalfa can protect against oxidative damage in fatty liver disease and hepatocyte injury[12,19], the underlying mechanisms remain unclear. We speculate that the antioxidant activity of APS might be attributable to a reduction in oxidative damage To investigate this possibility, an in vitro MEF model of oxidative stress induced by H2O2 was established. We characterized the antioxidative activities of APS in these cells and examined the signaling pathways involved
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