Hypoxia switches glucose depletion-induced necrosis to phosphoinositide 3-kinase/Akt-dependent apoptosis in A549 lung adenocarcinoma cells.
ABSTRACT In solid tumours, necrosis is commonly found in the core region in response to metabolic stress that results from oxygen and glucose depletion (OGD) due to insufficient vascularization and has been implicated in tumour progression. We have previously shown that metabolic stress due to glucose depletion (GD) induces necrosis and HMGB1 release through mitochondrial ROS production in A549 lung adenocarcinoma cells. In this study, we examined the effects of hypoxia on GD-induced necrosis and show that hypoxia prevented GD-induced mitochondrial ROS production, HMGB1 release, and necrosis and switched the cell death mode to apoptosis that is dependent on caspase-3 and -9. We further found that inhibition of ERK1/2 by U0126 abolished the effects of hypoxia to switch the cell death mode and to suppress mitochondrial ROS production, indicating an important role(s) of the ERK pathway in cell death mode determination. We also found that during OGD-induced apoptosis the prosurvival protein kinase Akt is activated and inhibition of Akt by the phosphoinositide 3-kinase (PI3K) inhibitors LY294002 and wortmannin prevent OGD-induced apoptosis, caspase-3 and -9 activation, and nuclear translocation of AIF and EndoG. Similar inhibitory effects of PI3K inhibitors were observed in A549 cells that underwent apoptosis when treated with GD in the presence of NAC (a general antioxidant) or catalase (a H(2)O(2) scavenger), or in the presence of active PKC by treatment with phorbol-12-myristate-13-acetate, indicating a crucial role(s) of the PI3K-Akt pathway in OGD-indcued apoptosis. In conclusion, our results demonstrate that hypoxia switches GD-induced necrosis to apoptosis and ERK1/2 and PI3K-Akt exert anti-necrotic and pro-apoptotic activities in the cell death, respectively.
- SourceAvailable from: Abhishek Kumar SinghACS Chemical Neuroscience 10/2012; · 3.87 Impact Factor
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ABSTRACT: An in vitro model of ischemic cerebral stroke [oxygen-glucose deprivation (OGD) for 6 h followed by 24 h reoxygenation (R)] with PC12 cells increases Ca(2+) influx by upregulating native L-type Ca(2+) channels and reactive oxygen species (ROS) generation. This reactive oxygen species generation and increase in intracellular Ca(2+) triggers the expression of hypoxic homeostasis transcription factors such as hypoxia induced factor-1 alpha (HIF-1α), Cav-beta 3 (Cav β3), signal transducer and activator of transcription 3 (STAT3), heat shock protein 27 (hsp-27), and cationic channel transient receptor potential melastatin 7 (TRPM7). OGD insulted PC12 cells were subjected to biologically safe doses (5, 10, and 25 μM) of trans-resveratrol in three different treatment groups: 24 h prior to OGD (pre-treatment); 24 h post OGD (post-treatment); and from 24 h before OGD to end of reoxygenation period (whole-treatment). Here, we demonstrated that OGD-R-induced neuronal injury/death is by reactive oxygen species generation, increase in intracellular calcium levels, and decrease in antioxidant defense enzymes. trans-Resveratrol increases the viability of OGD-R insulted PC12 cells, which was assessed by using MTT, NRU, and LDH release assay. In addition, trans-resveratrol significantly decreases reactive oxygen species generation, intracellular Ca(2+) levels, and hypoxia associated transcription factors and also increases the level of antioxidant defense enzymes. Our data shows that the whole-treatment group of trans-resveratrol is most efficient in decreasing hypoxia induced cell death through its antioxidant properties.ACS Chemical Neuroscience 02/2013; 4(2):285-94. · 3.87 Impact Factor
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ABSTRACT: The migration of hepatic stellate cells (HSCs) is essential to the hepatic fibrotic response, and recently High-mobility group box 1 (HMGB1) has been shown up-regulated during liver fibrosis. Nevertheless, whether HMGB1 can modulate the proliferation and migration of HSCs is poorly understood, as well as the involved intracellular signaling. In this study, we examined the effect of HMGB1 on proliferation, migration, pro-fibrotic function of HSCs and investigated whether toll-like family of receptor 4 (TLR4) dependent signal pathway is involved in the intracellular signaling regulation. METHODOLOGYPRINCIPAL FINDINGS: Modified transwell chamber system to mimic the space of Disse was used to evaluate the migration of human primary HSCs, and the protein expressions of related signal factors were evaluated by western blot. Cell proliferation was analyzed by MTT assay, the pro-fibrotic functions of HSCs by qRT-PCR and ELISA respectively. Recombinant human HMGB1 could significantly promote migration of HSCs under both haptotactic and chemotactic stimulation, especially the latter. Human TLR4 neutralizing antibody could markedly inhibit HMGB1-induced migration of HSCs. HMGB1 could enhance the phosphorylation of JNK and PI3K/Akt, and TLR4 neutralizing antibody inhibited HMGB1-enhanced phosphorylation of JNK and PI3K/Akt and activation of NF-κB. JNK inhibitor (SP600125) and PI3K inhibitor (LY 294002) significantly inhibited HMGB1-induced proliferation and migration of HSCs, and also reduced HMGB1-enhanced related collagen expressions and pro-fibrotic cytokines production. CONCLUSIONSSIGNIFICANCE: HMGB1 could significantly enhance migration of HSCs in vitro, and TLR4-dependent JNK and PI3K/Akt signal pathways are involved in the HMGB1-induced proliferation, migration and pro-fibrotic effects of HSCs, which indicates HMGB1 might be an effective target to treat liver fibrosis.PLoS ONE 01/2013; 8(5):e64373. · 3.73 Impact Factor