Xanthohumol-Induced Rat Glioma C6 Cells Death by Triggering Mitochondrial Stress

Abstract

AIM: To investigate the underlying mechanisms of xanthohumol (XN) on the proliferation inhibition and death of C6 glioma cells. METHODS: To determine the effects of XN on C6 cells, cell proliferation and mortality after XN treatment were assessed by SRB assay and trypan blue assay respectively. Apoptotic rates were evaluated by flowcytometry after Annexin V-FITC/PI double staining. The influence of XN on the activity of caspase-3 was determined by Western blot (WB); and nuclear transposition of apoptosis-inducing factor (AIF) was tested by immunocytochemistry and WB. By MitoSOXTM staining, the mitochondrial ROS were detected. Mitochondrial function was also tested by MTT assay (content of succinic dehydrogenase), flow cytometry (mitochondrial membrane potential (MMP)—JC-1 staining; mitochondrial abundance—mito-Tracker green), immunofluorescence (MMP—JC-1 staining; mitochondrial morphology—mito-Tracker green), WB (mitochondrial fusion-fission protein—OPA1, mfn2, and DRP1; mitophagy-related proteins—Pink1, Parkin, LC3B, and P62), and high-performance liquid chromatography (HPLC) (energy charge). Finally, mitochondrial protein homeostasis of C6 cells after XN treatment with and without LONP1 inhibitor bortezomib was investigated by trypan blue assay (proliferative activity and mortality) and WB (mitochondrial protease LONP1). All cell morphology images were taken by a Leica Microsystems microscope. RESULTS: XN could lead to proliferation inhibition and death of C6 cells in a time- and dose-dependent manner and induce apoptosis of C6 cells through the AIF pathway. After long incubation of XN, mitochondria of C6 cells were seriously impaired, and mitochondria had a diffuse morphology and mitochondrial ROS were increased. The content of succinic dehydrogenase per cell was significantly decreased after XN insults of 24, 48, and 72 h. The energy charge was weakened after XN insult of 24 h. Furthermore, the MMP and mitochondrial abundance were significantly decreased; the protein expression levels of OPA1, mfn2, and DRP1 were down-regulated; and the protein expression levels of Pink1, Parkin, LC3B-II/LC3B-I, and p62 were up-regulated in long XN incubation times (24, 48, and 72 h). XN incubation with bortezomib for 48 h resulted in lower proliferative activity and higher mortality of C6 cells and caused the cell to have visible vacuoles. Moreover, the protein expression levels of LONP1 was up-regulated gradually as XN treatment time increased. CONCLUSION: These data supported that XN could induce AIF pathway apoptosis of the rat glioma C6 cells by affecting the mitochondria.

Redox Activity of Flavonoids: Impact on Human Health, Therapeutics, and Chemical Safety

“…Similarly, findings by Yan et al demonstrated that intravenous administration of 3′-daidzein sulfonate sodium improved mitochondrial function after cerebral ischemia/reperfusion injury in rats through the reduction in mitochondrial swelling, elevation in mitochondrial membrane potential, increasing in GSH peroxidase and superoxide dismutase activities, and decreasing in mitochondrial malondialdehyde levels . Moreover, biochanin A plays an important role in preventing mitochondrial dysfunction by attenuating Aβ(25–35)-induced PC12 cell injury and sensitizing prostate cancer cells (DU145 and LNCap) to TRAIL-induced via a mitochondrial-dependent apoptosis pathway. , Another important flavonoid, xanthohumol, exhibited promising function in improving the mitochondrial function and preventing mitochondria form getting impaired with diffused morphology and increased ROS. , Furthermore, phloretin ameliorates arsenic trioxide-induced mitochondrial dysfunction in H9c2 cardiomyoblasts and reduces the mitochondrial trans-membrane potential through inducing the apoptosis of human hepatoma cells SMMC-7721 . In addition, some investigations reported that proanthocyanidins increases the activity of complex I and improves the mitochondrial dynamics through preventing irradiation-induced differentiation of human lung fibroblasts by ameliorating mitochondrial dysfunction .…”

“…160,161 Another important flavonoid, xanthohumol, exhibited promising function in improving the mitochondrial function and preventing mitochondria form getting impaired with diffused morphology and increased ROS. 37,162 Furthermore, phloretin ameliorates arsenic trioxide-induced mitochondrial dysfunction in H9c2 cardiomyoblasts 44 and reduces the mitochondrial trans-membrane potential through inducing the apoptosis of human hepatoma cells SMMC-7721. 163 In addition, some investigations reported that proanthocyanidins increases the activity of complex I and improves the mitochondrial dynamics through preventing irradiation-induced differentiation of human lung fibroblasts by ameliorating mitochondrial dysfunction.…”

The Protective Effects of α-Mangostin Attenuate Sodium Iodate-Induced Cytotoxicity and Oxidative Injury via Mediating SIRT-3 Inactivation via the PI3K/AKT/PGC-1α Pathway

“…The JC-1 staining assay is used to evaluate the function of mitochondria [ 29 ]. JC-1 monomers emit green fluorescence and indicate damaged mitochondria, while JC-1 polymers emit red fluorescence and indicate healthy mitochondria.…”

Cyanine-Functionalized 2,2′-Bipyridine Compounds for Photocatalytic Cancer Therapy

“…JC-1 emits red fluorescence when Δψ m is high and green fluorescence when the Δψ m is low. 31 As shown in Figure 3, the in-cell green fluorescence from JC-1 increased after bpyPCN+light treatment, indicating photoinduced depletion of Δψ m by bpyPCN. This change in the Δψ m might be a signature of cell apoptosis.…”