Abstract
Oxidative stress has been implicated in the pathogenesis of many diseases including chronic liver diseases. Nrf2 is a master transcriptional factor regulating the induction of cellular antioxidant defense systems. Here, the Nrf2-activating effect of the crude methanol extract of dried leaves of Pogostemon cablin Bentham was demonstrated by measuring the antioxidant response element (ARE)-driven luciferase activity and pachypodol, 4′,5-dihydroxy-3,3′,7-trimethoxyflavone, was isolated by bioactivity-guided fractionation and further separation using chromatographic techniques. To our knowledge, this is the first study to evaluate the antioxidant and cytoprotective effects of pachypodol in HepG2 cells as well as the underlying molecular mechanisms. Indeed, pachypodol protected HepG2 cells from cell death caused by tert-butylhydroperoxide-induced oxidative stress and also attenuated ROS production. The ability of pachypodol to activate Nrf2/ARE pathway was further confirmed by observing Nrf2 expression in nuclear fraction, mRNA levels of Nrf2 target antioxidants, and cellular glutathione content in HepG2 cells. Extracellular signal-regulated kinase (ERK) is one of the important kinases involved in Nrf2 activation. Pachypodol increased ERK phosphorylation and ERK inhibition by PD98059 totally abrogated the increase in ARE luciferase activity, nuclear Nrf2 accumulation and mRNA levels of antioxidant enzymes by pachypodol. In conclusion, pachypodol isolated from P. cablin can protect hepatocytes from oxidative injury, possibly mediated by enhancing endogenous antioxidant defense system through ERK-dependent Nrf2 activation.
Highlights
Reactive oxygen species (ROS) can play an important role in the physiological regulation of diverse cell functions, but excessively generated ROS can damage the structure and function of major cellular components, such as proteins, lipids and DNA [1]
The scheme for the extraction, fractionation and isolation of major active compounds from the crude methanol extract of dried leaves of P. cablin Bentham and HPLC chromatogram of crude extract are provided in Figure S1 and Figure 1a, respectively
We focused on the role of extracellular signal-regulated kinase (ERK), phosphoinositide 3-kinase (PI3K)/Akt, and AMP-activated protein kinase (AMPK) which have been known to be frequently involved in the activation of NF-E2-related factor-2 (Nrf2)/antioxidant response element (ARE) system stimulated by many phytochemicals [32,33,34,35]
Summary
Reactive oxygen species (ROS) can play an important role in the physiological regulation of diverse cell functions, but excessively generated ROS can damage the structure and function of major cellular components, such as proteins, lipids and DNA [1]. The importance of oxidative stress in the pathogenesis of various diseases suggests the potential of antioxidants for the prevention and treatment of these diseases. Upon exposure to oxidative stress or various antioxidants including phytochemicals, Nrf is dissociated from Nrf2-Kelch-like ECH-associated protein 1 (Keap1)-Cul complex in the cytoplasm and translocates into the nucleus [4]. Since the Nrf2/ARE signaling pathway is a key component of the coordinated induction of cellular antioxidant defense systems to maintain redox homeostasis [4], beneficial effects of pharmacological Nrf activators have been studied in many diseases in which oxidative stress is involved [6,7,8]
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