Effect of Tribulus terrestris saponins on proliferation of adipose-derived mesenchymal stem cells

Adipose-derived mesenchymal stem cells (ASCs) transplantation has shown great promise for treating various diseases; however, poor viability of transplanted ASCs because of oxidative stress has limited its therapeutic efficiency. Plant saponins are recently been reported to have antioxidant activity tested in various cancer cell lines. This study was designed to investigate the protective effects of Tribulus terrestris saponins (TTS) on the proliferation of ASCs. The cytotoxic activity of hydrogen peroxide (H₂O₂) was determined by treating ASCs with 100, 200, 300, 400, and 500 µM H₂O₂ for 2 hours. ASCs were treated with 6.25, 12.5, 25, 50, and 100 µg/mL concentrations of TTS for the proliferative experiment. To check the protective effect of TTS, experiments were designed in two ways. In one set, ASCs were pretreated with different concentrations of TTS for 2 hours and then apoptosis was induced by treating them with 400 µM H₂O₂ for next 2 hours, while in other set, ASCs were first treated with 400 µM H₂O₂ for 2 hours and subsequently with different concentrations of TTS for 24 hours. The vitality and proliferation potential of cells were detected by 3-(4,5-Dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The result of the current study shows that in response to stress-induced by H₂O₂ at concentration of 400 µM, ASCs underwent growth arrest and cell viability was reduced to half while treatment with TTS before and after H₂O₂ exposure significantly prevents premature apoptosis. The findings suggest that saponins may act as an effective protective agent against oxidative stress–induced ASCs apoptosis.     

Estradiol protects adipose tissue-derived stem cells against H(2) O(2) -induced toxicity

Oxidative stress is associated with various pathophysiological processes, including cell survival, adhesion, apoptosis, and cancer. In the present study, we aimed to evaluate the effects of H₂O₂‐induced toxicity on adipose tissue–derived stem cells (ADSCs) and whether 17β‐estradiol (E₂) has protective effects on these cells. ADSCs derived from adult Sprague–Dawley rats were pretreated with different doses of E₂ for 24 h and then exposed to 200 µM H₂O₂ for 4 h. Incubation of ADSCs with H₂O₂‐decreased cell viability in a concentration‐dependent fashion (p < 0.0001), whereas pretreatment of these cells with E₂ significantly reversed toxicity (p < 0.05), inhibited apoptotic changes, and decreased lipid peroxidation (p < 0.0005). Our findings suggest that E₂ protects ADSCs from oxidative‐induced cell death, and therefore, it may be used to improve the survival rate and regenerative capacity of stem cells. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:301–307, 2012; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21421     

Hydrogen peroxide modulates energy metabolism and oxidative stress in cultures of permanent human Müller cells MIO‐M1

In this study the influence of hydrogen peroxide (H₂O₂) on the redox state, NADH protein binding, and mitochondrial membrane potential in Müller cells is investigated. Cultures of permanent human Müller cells MIO‐M1 were exposed to H₂O₂ in 75 µM and 150 µM concentration for two hours. Fluorescence emission spectra and lifetimes were measured by two‐photon microscopy (excitation wavelength: 740 nm) at the mitochondria which were identified in the microscopic images by their fluorescence properties (spectra and intensity). Two hours of H₂O₂ exposure did not impair viability of MIO‐M1 cells in culture. Whereas the ratio of flavine‐ to NADH fluorescence intensity did not change under either H₂O₂ concentration, the mean lifetime was significantly different between controls, not exposed to H₂O₂, and the 150 µM H₂O₂ exposure (972 ± 63 ps vs. 1152 ± 64 ps, p = 0.014). One hour after cessation of the H₂O₂ exposure, the value retuned to that of the control (983 ± 36 ps). A hyperpolarization of the mitochondrial membrane under 150 µM H₂O₂ was found. These findings suggest a shift form free to protein‐bound NADH in mitochondria as well as a hyperpolarization of their inner membrane which could be related to an impairment of Müller cell function despite their preserved viability.

Exposure of human Müller cells to hydrogen peroxide for two hours results in a reversible change of protein binding of mitochondrial NADH upon unchanged redox ratio. The mitochondrial membrane potential is increased during exposure.     

Neuroprotective Effects of Phenolic and Carboxylic Acids on Oxidative Stress-Induced Toxicity in Human Neuroblastoma SH-SY5Y Cells

An increase in oxidative stress is a key factor responsible for neurotoxicity induction and cell death leading to neurodegenerative diseases including Parkinson’s and Alzheimer’s diseases. Plant phenolics exert diverse bioactivities i.e., antioxidant, anti-inflammatory, and neuroprotective effects. Herein, phenolic compounds, namely protocatechuic aldehyde (PCA) constituents of Hydnophytum formicarum Jack. including vanillic acid (VA) and trans-ferulic acid (FA) found in Spilanthes acmella Murr., were explored for anti-neurodegenerative properties using an in vitro model of oxidative stress-induced neuroblastoma SH-SY5Y cells. Exposure of the neuronal cells with H₂O₂ resulted in the decrease of cell viability, but increasing in the level of reactive oxygen species (ROS) together with morphological changes and inducing cellular apoptosis. SH-SY5Y cells pretreated with 5 µM of PCA, VA, and FA were able to attenuate cell death caused by H₂O₂-induced toxicity, as well as decreased ROS level and apoptotic cells after 24 h of treatment. Pretreated SH-SY5Y cells with phenolic compounds also helped to upregulate H₂O₂-induced depletion of the expressions of sirtuin-1 (SIRT1) and forkhead box O (FoxO) 3a as well as induce the levels of antioxidant (superoxide dismutase (SOD) 2 and catalase) and antiapoptotic B-cell lymphoma 2 (Bcl-2) proteins. The findings suggest that these phenolics might be promising compounds against neurodegeneration.     

Viridiflorol induces anti-neoplastic effects on breast,lung, and brain cancer cells through apoptosis

All active natural molecules are not fully exploited as therapeutic agents, causing delays in the advancement of anticancer drug discovery. Viridiflorol is a natural volatile element that may work as anti-cancer compound. We tested the anticancer properties of viridiflorol at different concentrations ranging from 0.03 to 300 μM in vitro on three cancer cells including breast (MCF-7), lung (A549) and brain (Daoy). The cancer cells responses were documented after treatment using MTT and Annexin V assays. Viridiflorol showed cytotoxic effects against all tested cell lines, reducing cell viability in a concentration-dependent manner with variable IC₅₀ values. Daoy and A549 cell lines were more sensitive to viridiflorol when compared with temozolomide and doxorubicin, respectively. Viridiflorol demonstrated the highest anticancer activity against the Daoy cells with an estimated IC₅₀ of 0.1 µM followed by MCF-7 at 10 µM, and A549 at 30 µM. In addition, upon exposure to concentrations ranging from 30 µM to 300 µM of viridiflorol, early and late apoptotic cell death was induced in a concentration dependent manner in Daoy (55.8%-72.1%), MCF-7 (36.2%-72.7%) and A459 (35%-98.9%) cell lines, respectively. In conclusion, viridiflorol demonstrates cytotoxic and apoptotic ability in three different cancer cell lines (brain, breast and lung).     

Suppression of histamine-induced relaxation of rat aorta and calcium signaling in endothelial cells by two-pore channel blocker trans-NED19 and hydrogen peroxide

The blocker of two-pore channels trans-NED19 and hydrogen peroxide were found to inhibit histamine-induced relaxation of rat aorta. The degree of inhibition depended on histamine concentration. The relaxation in response to 1 µM histamine of rat aorta preconstricted with 30 mM KCl, serotonin, or endothelin-1, was completely abolished by 30 µM trans-NED19. On the other hand, trans-NED19 decreased the relaxation of the aorta in the presence of 10 µM histamine only by 2.1to 2.4-fold, and there was almost no inhibition by trans-NED19 of the relaxation induced by 100 µM histamine. Relaxation of preconstricted with serotonin aorta in response to 10 and 100 µM histamine was reduced by hydrogen peroxide (200 µM) by 10and 2.5-fold, respectively. Suppression of aorta relaxation by trans-NED19 and H₂O₂ correlated with their inhibitory effect on the histamine-induced increase in the cytoplasmic free calcium concentration in human umbilical vein endothelial cells. With the use of a fluorescent probe LysoTracker, the cis-NED19 binding sites were demonstrated to be localized in endolysosomes of the endothelial cells. These data indicate that twopore calcium channels participate in the histamine-induced endothelium-dependent relaxation of rat aorta. Furthermore, their functional role is exhibited much more clearly at low histamine concentrations. We suggest that hydrogen peroxide evokes depletion of intracellular calcium depots thereby suppressing the response to histamine.     

Hydrogen peroxide in the reactions of cancer cells to cisplatin

BackgroundHydrogen peroxide (H₂O₂) is thought to be one of the key components involved in the responses of tumor cells to chemotherapy. The aim of this study was to reveal the pathways and the phases of cisplatin-induced cell death that are characterized by changes of H₂O₂ level.MethodsThe genetically encoded cytosolic fluorescent sensor HyPer2 was used for flow cytometric analysis of the cisplatin-induced changes in H₂O₂ level in HeLa Kyoto cells. Using a vital dye and the apoptotic markers PE Annexin V or TMRE the pathways and stages of cell death were investigated simultaneously with HyPer2 response. The H₂O₂ level was studied separately in viable and early apoptotic cells after 12, 18, 24 h's incubation with cisplatin at several concentrations with or without the scavenger of reactive oxygen species NAC.ResultsCisplatin causes dose- and time-dependent increase of H₂O₂ level in TMRE-positive and PE Annexin V-negative cancer cells. The scavenging of ROS by NAC decreased the H₂O₂ level and restored cell viability.ConclusionН₂О₂ generation begins in cells that have already lost mitochondrial membrane potential but have not yet externalized phosphatidylserine. Prevention of apoptosis by NAC confirmed the role of H₂O₂ in apoptosis induction.General significanceThis is the first time that the sensor HyPer2 has been used in parallel with apoptotic markers and vital dye to demonstrate the role of H₂O₂ in different stages and types of tumor cell death under chemotherapeutic action.     

Fibroblast growth factor-8 inhibits oxidative stress-induced apoptosis in H9c2 cells

Fibroblast growth factors (FGFs) comprise a large family of signaling molecules that involve cell patterning, mobilization, differentiation, and proliferation. Various FGFs, including FGF-1, FGF-2, and FGF-5, have been shown to play a role in cytoprotection during adverse cardiac events; however, whether FGF-8 is a cytoprotective remains unclear. The current study was designed to evaluate the effect of FGF-8 treatment on oxidative stress-induced apoptosis in H9c2 cells. Cells were divided into three groups: control, H₂O₂ (400 µm H₂O₂), and H₂O₂ + FGF-8 (4 ng/ml FGF-8). Our results suggest apoptosis was significantly (p < 0.05) enhanced in the H₂O₂ group relative to control. Moreover, a significant (p < 0.05) decline in apoptosis was observed in the H₂O₂ + FGF-8 group compared to H₂O₂-treated cells as evidenced by TUNEL staining, a cell death detection ELISA, and cell viability. Levels of downstream apoptotic mediators, caspase-3 and caspase-9, were significantly (p < 0.05) upregulated following H₂O₂ treatment but were abrogated following FGF-8 application. Expression levels of Forkhead box protein O1 (FoxO-1), MnSOD, catalase, pAKT, and p-mTOR were significantly (p < 0.05) reduced in the H₂O₂ group (p < 0.05). Notably, these levels were significantly (p < 0.05) reversed following FGF-8 treatment. Our data, for the first time, suggest FGF-8 is an anti-apoptotic mediator in oxidative-stressed H9c2 cells. Furthermore, our data demonstrate that apoptotic inhibition by FGF-8 is consequent to FoxO-1 oxidative detoxification as well as augmentation to the PI3K/AKT cell survival pathway.     

Neuroprotective effects of corn silk maysin via inhibition of H2O2-induced apoptotic cell death in SK-N-MC cells

Aims

Neuroprotective effects of maysin, which is a flavone glycoside that was isolated from the corn silk (CS, Zea mays L.) of a Korean hybrid corn Kwangpyeongok, against oxidative stress (H₂O₂)-induced apoptotic cell death of human neuroblastoma SK-N-MC cells were investigated.

Main methods

Maysin cytotoxicity was determined by measuring cell viability using MTT and lactate dehydrogenase (LDH) assays. Intracellular reactive oxygen species (ROS) were measured using a 2,7-dichlorofluorescein diacetate (DCF-DA) assay. Apoptotic cell death was monitored by annexin V-FITC/PI double staining and by a TUNEL assay. Antioxidant enzyme mRNA levels were determined by real-time PCR. The cleavage of poly (ADP-ribose) polymerase (PARP) was measured by western blotting.

Key findings

Maysin pretreatment reduced the cytotoxic effect of H₂O₂ on SK-N-MC cells, as shown by the increase in cell viability and by reduced LDH release. Maysin pretreatment also dose-dependently reduced the intracellular ROS level and inhibited PARP cleavage. In addition, DNA damage and H₂O₂-induced apoptotic cell death were significantly attenuated by maysin pretreatment. Moreover, maysin pretreatment (5–50 μg/ml) for 2 h significantly and dose-dependently increased the mRNA levels of antioxidant enzymes (CAT, GPx-1, SOD-1, SOD-2 and HO-1) in H₂O₂ (200 μM)-insulted cells.

Significance

These results suggest that CS maysin has neuroprotective effects against oxidative stress (H₂O₂)-induced apoptotic death of human brain SK-N-MC cells through its antioxidative action. This report is the first regarding neuroprotective health benefits of corn silk maysin by its anti-apoptotic action and by triggering the expression of intracellular antioxidant enzyme systems in SK-N-MC cells.     

Madecassoside,a triterpenoid saponin isolated from Centella asiatica herbs,protects endothelial cells against oxidative stress

This study aimed to investigate the effect of madecassoside against oxidative stress‐induced injury of endothelial cells. Hydrogen peroxide (H₂O₂, 500 µmol/L) was employed as an inducer of oxidative stress in human umbilical vein endothelial cells (HUVECs). Cell apoptosis was detected by Hoechst 33258 staining and flow cytometry. Caspase‐3 activity and mitochondria membrane potential were further examined. As a result, madecassoside (10, 30, 100 µmol/L) could reverse morphological changes, elevate cell viability, increase glutathione levels, and decrease lactate dehydrogenase and malondialdehyde levels caused by H₂O₂ in a concentration‐dependent manner. It attenuated apoptosis, preventing the activation of caspase‐3 and the loss of mitochondria membrane potential, as well as the phosphorylation of p38 mitogen‐activated protein kinase (MAPK) in HUVECs. These data suggested that madecassoside could protect HUVECs from oxidative injury, which was probably achieved by inhibiting cell apoptosis via protection of mitochondria membranes and downregulation of the activation of caspase‐3 and p38 MAPK. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:399–406, 2012; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21434     

Role of PTHrP in human intestinal Caco-2 cell response to oxidative stress

We have previously demonstrated that parathyroid hormone (PTH) induces apoptosis in human colon adenocarcinoma Caco-2 cells but the effects of its tumoral analog PTH-related peptide (PTHrP) in this cell line are still unknown. In the present work we investigated whether PTHrP, as PTH, is able to induce Caco-2 cell apoptosis or if it exerts protective effects under apoptotic conditions. Using Caco-2 cells cultured under serum deprivation in the presence or absence of PTHrP we demonstrated that, differently to PTH, its analog employed at the same concentration (10^? 8 M) is not a pro-apoptotic hormone. Cells were exposed to an oxidative insult in the form of hydrogen peroxide to induce apoptosis, which leads to a 50% loss of cell viability determined by MTS assay, morphological changes observed under fluorescence microscopy and Western blot analysis. Herein we demonstrate, for the first time, that pre-treatment with PTHrP prior to H₂O₂ incubation, prevents cell death induced by the apoptotic inductor; and using specific inhibitors we evidenced that protein kinase B (AKT), extracellular signal-regulated kinase 1/2 (ERK1/2), c-Jun N-terminal kinase 1/2 (JNK1/2) and p38 mitogen-activated protein kinase (MAPK) mediate this anti-apoptotic effect. Also, we found that PTHrP decreases the pro-apoptotic protein BAX levels and increases the protein expression of the anti-apoptotic HSP27. Immunoblot analysis revealed that H₂O₂ increases the phosphorylation levels of AKT and MAPKs, exhibiting a cellular defense response; and consequently increases phospho-BAD levels. The H₂O₂-induced activation of protein kinases is reverted when cells are pre-treated with PTHrP. Altogether these results evidence a protective effect of PTHrP under apoptotic conditions in intestinal cells, which may be mediated by AKT and MAPKs.     

Oxidative stress-induced DNA damage and cell cycle regulation in B65 dopaminergic cell line

Reactive oxygen species and oxidative stress are associated with neuronal cell death in many neurodegenerative conditions. However, the exact molecular mechanisms triggered by oxidative stress in neurodegeneration are still unclear. This study used the B65 rat neuroblastoma cell line as a model to study the molecular events that occur after H₂O₂ treatment. Treatment of B65 cells with H₂O₂ rapidly up-regulated the DNA damage pathway involved in double-strand breakage. Subsequently, proteins involved in p53 regulation, such as sirtuin 1 and STAT1, were modified. In addition, H₂O₂ treatment altered the pattern of cell cycle protein expression. Specifically, a decrease was found in the expression of cyclin D1, cdk4 and surprisingly the levels of cyclin A and the retinoblastoma protein phosphorylated at ser780 were increased. Furthermore, this study shows that pre-treatment of B65 cells with 50 µM trolox confers almost total protection against apoptotic cell death and restores the cell cycle. Likewise, the increase in retinoblastoma phosphorylation was attenuated by KU-55993, a selective ATM inhibitor, and also by trolox. These observations indicate that DNA damage and oxidative stress are responsible for cell cycle regulation. In summary, this study describes the molecular mechanisms involved in cell cycle alterations induced by oxidative stress in B65 cells. These findings highlight the relevance of ATM in the regulation of cell cycle after oxidative stress.     

Biphasic regulation of H2O2 on angiogenesis implicated NADPH oxidase

ROS (reactive oxygen species) take an important signalling role in angiogenesis. Although there are several ways to produce ROS in cells, multicomponent non‐phagocytic NADPH oxidase is an important source of ROS that contribute to angiogenesis. In the present work, we examined the effects of H₂O₂ on angiogenesis including proliferation and migration in HUVECs (human umbilical vein endothelial cells), new vessel formation in chicken embryo CAM (chorioallantoic membrane) and endothelial cell apoptosis, which is closely related to anti‐angiogenesis. Our results showed that H₂O₂ dose‐dependently increased the generation of O₂ ^? (superoxide anion) in HUVECs, which was suppressed by DPI (diphenylene iodonium) and APO (apocynin), two inhibitors of NADPH oxidase. H₂O₂ at low concentrations (10 µM) stimulated cell proliferation and migration, but at higher concentrations, inhibited both. Similarly, H₂O₂ at 4 nmol/cm² strongly induced new vessel formation in CAM, while it suppressed at high concentrations (higher than 4 nmol/cm²). Also, H₂O₂ (200～500 µM) could stimulate apoptosis in HUVECs. All the effects of H₂O₂ on angiogenesis could be suppressed by NADPH oxidase inhibitors, which suggests that NADPH oxidase acts downstream of H₂O₂ to produce O₂ ^? and then to regulate angiogenesis. In summary, our results suggest that H₂O₂ as well as O₂ ^? mediated by NADPH oxidase have biphasic effects on angiogenesis in vitro and in vivo.     

The role of insulin against hydrogen peroxide-induced oxidative damages in differentiated SH-SY5Y cells

Abstract

Exogenous hydrogen peroxide (H₂O₂) can easily penetrate into biological membranes and enhance the formation of other reactive oxygen species (ROS). In the present study, we have investigated the neuroprotective effects of insulin on H₂O₂-induced toxicity of retinoic acid (RA)-differentiated SH-SY5Y cells. To measure the changes in the cell viability of SH-SY5Y cells at different concentrations of H₂O₂ for 24?h, a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT)-based assay was used and a 100?µM H₂O₂ was selected to establish a model of H₂O₂-induced oxidative stress. Further assays showed that 24?h of 100?µM H₂O₂-induced significant changes in the levels of lactate dehydrogenase (LDH), nitric oxide (NO), ROS, and calcium ion (Ca²⁺) in neuronal cells, but insulin can effectively diminish the H₂O₂-induced oxidative damages to these cells. Moreover, cells treated with insulin increased H₂O₂-induced suppression of glutathione levels and exerted an apparent suppressive effect on oxidative products. The results of insulin treatment with SH-SY5Y cells increased the Bcl-2 levels and decreased the Akt levels. The treatment of insulin had played a protective effect on H₂O₂-induced oxidative stress related to the Akt/Bcl-2 pathways.     

Deferoxamine Preconditioning of Neural-Like Cells Derived from Human Wharton’s Jelly Mesenchymal Stem Cells as a Strategy to Promote Their Tolerance and Therapeutic Potential: An In Vitro Study

Transplantation of neural-like cells is considered as a promising therapeutic strategy developed for neurodegenerative disease in particular for ischemic stroke. Since cell survival is a major concern following cell implantation, a number of studies have underlined the protective effects of preconditioning with hypoxia or hypoxia mimetic pharmacological agents such as deferoxamine (DFO), induced by activation of hypoxia inducible factor-1 (HIF-1) and its target genes. The present study has investigated the effects of DFO preconditioning on some factors involved in cell survival, angiogenesis, and neurogenesis of neural-like cells derived from human Wharton’s jelly mesenchymal stem cells (HWJ-MSCs) in presence of hydrogen peroxide (H₂O₂). HWJ-MSCs were differentiated toward neural-like cells for 14 days and neural cell markers were identified using immunocytochemistry. HWJ-MSC-derived neural-like cells were then treated with 100 µM DFO, as a known hypoxia mimetic agent for 48 h. mRNA and protein expression of HIF-1 target genes including brain-derived neurotrophic factors (BDNF) and vascular endothelial growth factor (VEGF) significantly increased using RT-PCR and Western blotting which were reversed by HIF-1α inhibitor, while, gene expression of Akt-1, Bcl-2, and Bax did not change significantly but pAkt-1 was up-regulated as compared to poor DFO group. However, addition of H₂O₂ to DFO-treated cells resulted in higher resistance to H₂O₂-induced cell death. Western blotting analysis also showed significant up-regulation of HIF-1α, BDNF, VEGF, and pAkt-1, and decrease of Bax/Bcl-2 ratio as compared to poor DFO. These results may suggest that DFO preconditioning of HWJ-MSC-derived neural-like cells improves their tolerance and therapeutic potential and might be considered as a valuable strategy to improve cell therapy.     

Prooxidative effects of green tea polyphenol (−)-epigallocatethin-3-gallate on the HIT-T15 pancreatic beta cell line

Epigallocatechin-3-gallate (EGCG) is the main polyphenolic constituent in green tea and is believed to function as an antioxidant. However, increasing evidence indicates that EGCG produces reactive oxygen species (ROS) and subsequent cell death. In this study, we investigated the prooxidative effects of EGCG on the HIT-T15 pancreatic beta cell line. Dose-dependent cell viability was monitored with the cell counting kit-8 assay, while the induction of apoptosis was analyzed by a cell death ELISA kit and comet assay. Extracellular H₂O₂ was determined using the Amplex Red Hydrogen Peroxide Assay Kit. Intracellular oxidative stress was measured by fluorometric analysis of 2′,7′-dichlorofluorescin (DCFH) oxidation using DCFH diacetate (DA) as the probe. Treatment with EGCG (5–100 μM) decreased the viability of pancreatic beta cells, caused concomitant increases in apoptotic cell death, and increased the production of H₂O₂ and ROS. Catalase, the iron-chelating agent diethylenetriaminepentaacetic acid, and the Fe(II)-specific chelator o-phenanthroline all suppressed the effects of EGCG, indicating the involvement of both H₂O₂ and Fe(II) in the mechanism of action of EGCG. The antioxidant N-acetyl-cysteine and alpha-lipoic acid also suppressed the effects of EGCG. Furthermore, EGCG did not scavenge exogenous H₂O₂, but rather, it synergistically increased H₂O₂-induced oxidative cell damage in pancreatic beta cells. Together, these findings suggest that in the HIT-T15 pancreatic beta cell line, EGCG mediated the generation of H₂O₂, triggering Fe(II)-dependent formation of a highly toxic radical that in turn induced oxidative cell damage.     

Peroxisome proliferator-activated receptor-α activation protects against endoplasmic reticulum stress-induced HepG2 cell apoptosis

Live ischemia–reperfusion injury is associated with endoplasmic reticulum (ER) stress-induced apoptosis. Activation of peroxisome proliferator-activated receptor-α (PPARα) may inhibit hepatocyte apoptosis induced by oxidative stress and protect against liver injury. This study aimed to investigate the effects of PPARα activation, through a specific agonist, on ER stress-induced apoptosis in human liver hepatocellular carcinoma (HepG2) cells. HepG2 cells were challenged with H₂O₂ and treated with WY14643, a selective PPARα agonist, in the presence or absence of the PPARα antagonist of MK886. Cell viable assay (MTT) and immunostaining were used to evaluate cell viability. The level of apoptotic cell death was quantified through Annexin V/PI staining. Alanine aminotransferase, asparatate aminotransferase, and malondialdehyde levels were measured to determine the presence of cellular injury and oxidative stress. RT-PCR and Western blot analysis were used to detect mRNA and protein expression of PPARα, BiP, and CHOP. Immunofluorescence was utilized to determine the intracellular localization of CHOP. H₂O₂ and MK886 both reduced the viability of HepG2 cells, increased oxidative stress and apoptosis, up-regulated the BiP and CHOP expression, and induced CHOP translocation from the cytoplasm to the nucleus. Compared with cells treated with H₂O₂ alone, pre-administration of WY14643 increased cell viability, attenuated apoptosis, improved cell function, down-regulated BiP and CHOP expression and inhibited CHOP translocation. The effects of WY14643 were completely abolished using the MK886 antagonist. PPARα activation protects against H₂O₂-induced HepG2 cell apoptosis. The underlying mechanisms may be associated with its activation to suppress excessive ER stress.     

Effect of lycopene on As2O3 induced oxidative stress in SH-SY5Y cells

It is known that oxidative stress may cause neuronal injury and several experimental models showed that As₂O₃ exposure causes oxidative stress. Lycopene, a carotenoid, has been shown to have protective effect in neurological disease models due to antioxidant activity, but its effect on As₂O₃-induced neurotoxicity is not identified yet. The aim of this study is to investigate the effects of lycopene on As₂O₃-induced neuronal damage and the related mechanisms. Cell viability was determined by the MTT assay. Lycopene was administrated with different concentrations (2, 4, 6 and 8 µM) one hour before 2 µM As₂O₃ exposure in SH-SY5Y human neuroblastoma cells. The anti-oxidant effect of lycopene was determined by measuring superoxide dismutase (SOD), catalase (CAT) hydrogen peroxide (H₂O₂), malondialdehyde (MDA), total antioxidant status (TAS) and total oxidant status (TOS). MTT results and LDH cytotoxicity analyses showed that pretreatment with 8 µM lycopene significantly improved the toxicity due to As₂O₃ exposure in SH?SY5Y neuroblastoma cells. Pretreatment with lycopene significantly increased the activities of anti?oxidative enzymes as well as total antioxidant status and decreased total oxidative status in As₂O₃ exposed cells. The results of this study indicate that lycopene may be a potent neuroprotective against oxidative stress and could be used to prevent neuronal injury or death in several neurological diseases.