Puerarin Inhibits FUNDC1-mediated Mitochondrial Autophagy and CSE-induced Apoptosis of Human Bronchial Epithelial Cells by Activating the PI3K/AKT/MTOR Signaling Pathway

Abstract

Background: Growing evidence suggests that the pathogenesis of chronic obstructive pulmonary disease (COPD) is related to mitochondrial autophagy mediated by FUN14 domain protein 1 (FUNDC1). Recent studies have shown that puerarin has protective effects against excessive oxidative damage in cells. We hypothesized that puerarin might be involved in the progression of COPD induced by cigarette smoke extract (CSE) by regulating FUNDC1-mediated mitochondrial autophagy.Material/Methods: Different concentrations of puerarin were used to intervene in CSE-treated human bronchial epithelial cells (HBECs). MTT assay and flow cytometry were used to detect cell activity and apoptosis. The mitochondrial membrane potential (MMP) level, reactive oxygen species (ROS) content and ATP content were detected by flow cytometry or kits; Western Blotting was used to detect mitochondrial autophagy-related proteins expression such as DRP1, FUNDC1, PINK1, Parkin. Next, protein phosphatase inhibitor (PH0321) was used to inhibit the phosphorylation of FUNDC1, the changes of MMP, ROS, ATP and mitochondrial autophagy related proteins were detected. Furthermore, mitochondrial autophagy inhibitors (Mdivi) were used to block autophagy, and then the changes of mitochondrial autophagy, cell activity and apoptosis were detected, respectively. Finally, the changes of PI3K/AKT/mTOR signaling pathway-related proteins were detected by Western Blotting.Results: Puerarin reversed cell activity decrease and apoptosis increase of CSE-induced HBECs, and down-regulated apoptosis-related proteins expression. Puerarin up-regulated MMP level and ATP content in CSE-induced HBECs, and down-regulated ROS content and mitochondrial autophagy-related proteins expression. PH0321 promotes mitochondrial autophagy by promoting the dephosphorylation of FUNDC1, puerarin inhibits the dephosphorylation of FUNDC1, up-regulated the MMP level and ATP content of CSE-induced HBECs, reduces ROS content and mitochondrial autophagy-related proteins expression. Compared with Mdivi group, the protein expression changes of PINK1 and Parkin in puerarin-treated group were not significantly different, and the cell activity decrease and apoptosis increase of CSE-induced HBECs were also significantly reversed by puerarin, which was not significantly different from Mdivi group. Puerarin significantly upregulated the protein expression of p-PI3K, p-AKT and p-mTOR, suggesting that puerarin may participate in the progression of COPD by activating the PI3K/AKT/mTOR signaling pathway.Conclusions: Puerarin inhibits FUNDC1-mediated mitochondrial autophagy and CSE-induced apoptosis of HBECs by activating the PI3K/AKT/mTOR signaling pathway.

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“…Azaleatin is also a avonoid with a wide range of pharmacological effects, inhibiting glutamate-induced reactive oxygen species expression, apoptosis, and mitochondrial damage in HT3 cells [26] . Puerarin can reduce CSEinduced apoptosis in human bronchial epithelial cells by modulating the PI3K/Akt/mTOR signaling pathway [50] .…”

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“…The PI3K/AKT/mTOR pathway is the most important mechanism of autophagy in cells, and regulating this signaling pathway can affect the occurrence and development of tumors [29]. Wang et al found that blocking the PI3K/AKT/mTOR pathway inhibited the progression of cervical cancer [30].…”