Thymoquinone Induces Cell Death in Human Squamous Carcinoma Cells via Caspase Activation-Dependent Apoptosis and LC3-II Activation-Dependent Autophagy

Abstract

BackgroundThymoquinone (TQ), an active component of Nigella sativa or black cumin, elicits cytotoxic effects on various cancer cell lines. However, the anti-cancer effects of TQ on head and neck squamous cell carcinoma (HNSCC) remain unclear.Methodology/Principal FindingsIn this study, TQ elicited a strong cytotoxic effect on SASVO3, a highly malignant HNSCC cell line. The mechanisms of this cytotoxic effect were concentration dependent. TQ also induced apoptotic cell death in SASVO3 cells as indicated by an increase in Bax expression and caspase-9 activation. Apoptosis was possibly caspase-9 dependent because the exposure of cells to a caspase-9 inhibitor partially prevented cell death. The exposed cells also showed increased levels of autophagic vacuoles and LC3-II proteins, which are specific autophagy markers. Cell viability assay results further revealed that bafilomycin-A1, an autophagy inhibitor, enhanced TQ cytotoxicity; by comparison, Annexin V and propidium-iodide staining assay results showed that this inhibitor did not promote apoptosis. TQ treatment also increased the accumulation of autophagosomes. Using a lentivirus-shRNA system for LC3 silencing, we found that cell viability was eradicated in autophagy-defective cells. An in vivo BALB/c nude mouse xenograft model further showed that TQ administered by oral gavage reduced tumor growth via induced autophagy and apoptosis.ConclusionsThese findings indicated that TQ induced cell death in oral cancer cells via two distinct anti-neoplastic activities that can induce apoptosis and autophagy. Therefore, TQ is a promising candidate in phytochemical-based, mechanistic, and pathway-targeted cancer prevention strategies.

Thymoquinone chemosensitizes colon cancer cells through inhibition of NF-κB

“…In the previous decade, the antitumor activity of TQ has been investigated in a number of studies (6)(7)(8). TQ was observed to induce antitumor effects in several types of cancer, including breast (9), lung (10), multiple myeloma (11), pancreatic (12), cervical (13), colon (14) and prostate cancer (15), as well as squamous (16) and hepatocellular carcinoma (17), acute lymphoblastic leukemia (18), glioblastoma (19), osteosarcoma (20), neuroblastoma (21), bladder (22), gastric (23) and ovarian cancer (24). Although the effect of TQ has been investigated in numerous types of cancer, the molecular mechanisms underlying its action remain to be elucidated.…”

Capsaicin protects endothelial cells and macrophage against oxidized low-density lipoprotein-induced injury by direct antioxidant action

“…The inhibitory effects of capsaicin on ApoB fragmentation were examined by SDS-PAGE. The ApoB band (arrow) in native LDL disappeared because of fragmentation caused by incubation with CuSO 4 for 4 h. The presence of capsaicin (10,20,30,40, and 50 lM) significantly reversed copper-induced ApoB fragmentation (Fig. 1A).…”

“…Human plasma was obtained from the Taichung Blood Bank, and LDL was isolated using sequential ultracentrifugation [20]. LDL (q = 1.019-1.210 g/mL) in KBr solution containing 30 mM EDTA was stored at 4°C in a sterile, dark environment and used within 3 d. Immediately before oxidation tests, LDL was separated from EDTA and diffusible low molecular mass compounds by gel filtration on PD-10 gel (Pharmacia, St-Quentin, France) in 10 mM PBS (pH 7.4).…”

“…In brief, 1 Â 10 5 cells were fixed in 1 mL of 70% ethanol for 30 min, and suspended in 1 mL of propidium iodide (PI) solution (25 lg/mL PI, 0.1 mM EDTA, and 10 lg/mL RNase A in PBS) for 30 min in the dark. Cell cycle distribution was then analyzed by flow cytometry analysis using CellQuest software (Becton-Dickinson, CA, USA) [20].…”

“…JC-1 exhibits potential-dependent accumulation in the mitochondria, as indicated by the fluorescence emission shift from 530 to 590 nm. After treatment of oxLDL (200 lg/mL) for 16 h in the presence or absence of capsaicin (10,20,30,40, and 50 lM), cells (5 Â 10 4 cells/24-well plates) were rinsed with M199 followed by the addition of JC-1 (5 lM). After incubation at 37°C for 20 min, cells were examined under a fluorescent microscope [21].…”