Phenylethanoid Glycosides From Callicarpa kwangtungensis Chun Attenuate TNF-α-Induced Cell Damage by Inhibiting NF-κB Pathway and Enhancing Nrf2 Pathway in A549 Cells.

Background: Acute lung injury (ALI) is a complicated and severe lung disease, which is often characterized by acute inflammation. Poliumoside (POL), acteoside (ACT) and forsythiaside B (FTB) are phenylethanoid glycosides (PGs) with strong antioxidant, anti-inflammatory, and anti-apoptotic properties, which are extracted from Callicarpa kwangtungensis Chun (CK). The aim of this study was to investigate the protective effects of POL, ACT, and FTB against TNF-α-induced damage using an ALI cell model and explore their potential mechanisms. Methods and Results: MTT method was used to measure cell viability. Flow cytometry was used for detecting the apoptosis rate. Reactive oxygen species (ROS) activity was determined using fluorescence microscope. The expression of mRNA in apoptosis-related genes (Caspase 3, Caspase 8, and Caspase 9) were tested by qPCR. The effects of POL, ACT, FTB on the activities of nuclear factor erythroid-2 related factor 2 (Nrf2), nuclear factor kappa-B (NF-κB) and the expression of their downstream genes were assessed by western blotting and RT-PCR in A549 cells. In the current study, POL, ACT, and FTB dose-dependently attenuated TNF-α-induced IL-1ß, IL-6 and IL-8 production, cell apoptosis, the expression of apoptosis-related genes (Caspase 3, Caspase 8, and Caspase 9) and ROS activity. POL, ACT, and FTB not only increased in the mRNA levels of antioxidative enzymes NADPH quinone oxidoreductase (NQO1), glutamate cysteine ligase catalytic subunit (GCLC), heme oxygenase (HO-1), but also decreased the mRNA levels of IL-1ß, IL-6 and IL-8. Furthermore, they upregulated the expression of Keap1 and enhanced the activation of Nrf2, while decreased the expression of phosphor-IκBα (p-IκBα) and nuclear p65. In addition, no significant changes were observed in anti-inflammatory and antioxidant effects of POL, ACT, FTB following Nrf2 and NF-κB p65 knockdown. Conclusion: Our study revealed that POL, ACT, and FTB alleviated oxidative damage and lung inflammation of TNF-α-induced ALI cell model through regulating the Nrf2 and NF-κB pathways.

miR­29b suppresses proliferation and induces apoptosis of hepatocellular carcinoma ascites H22 cells via regulating TGF­ß1 and p53 signaling pathway.

MicroRNA (miR)­29b is a key tumor regulator. It can inhibit tumor cell proliferation, induce apoptosis, suppress tumor invasion and migration, thus delaying tumor progression. Our previous studies revealed an increased level of miR­29b in hepatoma 22 (H22) cells in ascites tumor­bearing mice. The present study investigated the effect of miR­29b on proliferation and apoptosis of hepatocellular carcinoma ascites H22 cells and its association with the transforming growth factor­ß1 (TGF­ß1) signaling pathway and p53­mediated apoptotic pathway. Briefly, H22 cells were transfected with miR­29b­3p (hereinafter referred to as miR­29b) mimic or miR­29b inhibitor. MTS cell proliferation assay and flow cytometry were used to analyze cell viability and apoptosis. The expression change of the TGF­ß1 signaling pathway and p53­mediated apoptotic pathway were detected by reverse transcription­quantitative PCR, western blotting and immunofluorescence. Furthermore, cells were treated with exogenous TGF­ß1 and TGF­ß1 small interfering RNA to evaluate the crosstalk between TGF­ß1 and p53 under miR­29b regulation. The overexpression of miR­29b decreased cell viability, increased cell apoptosis, activated the TGF­ß1 signaling pathway and p53­mediated apoptotic pathway. Conversely, these effects were reversed by the miR­29b inhibitor. Moreover, the effect of miR­29b mimic was further increased after treating cells with exogenous TGF­ß1. The activation of the TGF­ß1 signaling pathway and p53­mediated apoptotic pathway induced by miR­29b overexpression were reversed by TGF­ß1 inhibition. In summary, these data indicated that miR­29b has an important role in proliferation and apoptosis of H22 cells by regulating the TGF­ß1 signaling pathway, the p53­dependent apoptotic pathway, and the crosstalk between TGF­ß1 and p53.