4β‐Hydroxywithanolide E selectively induces oxidative DNA damage for selective killing of oral cancer cells

Reactive oxygen species (ROS) induction had been previously reported in 4β‐hydroxywithanolide (4βHWE)‐induced selective killing of oral cancer cells, but the mechanism involving ROS and the DNA damage effect remain unclear. This study explores the role of ROS and oxidative DNA damage of 4βHWE in the selective killing of oral cancer cells. Changes in cell viability, morphology, ROS, DNA double strand break (DSB) signaling (γH2AX foci in immunofluorescence and DSB signaling in western blotting), and oxidative DNA damage (8‐oxo‐2′deoxyguanosine [8‐oxodG]) were detected in 4βHWE‐treated oral cancer (Ca9‐22) and/or normal (HGF‐1) cells. 4βHWE decreased cell viability, changed cell morphology and induced ROS generation in oral cancer cells rather than oral normal cells, which were recovered by a free radical scavenger N‐acetylcysteine (NAC). For immunofluorescence, 4βHWE also accumulated more of the DSB marker, γH2AX foci, in oral cancer cells than in oral normal cells. For western blotting, DSB signaling proteins such as γH2AX and MRN complex (MRE11, RAD50, and NBS1) were overexpressed in 4βHWE‐treated oral cancer cells in different concentrations and treatment time. In the formamidopyrimidine‐DNA glycolyase (Fpg)‐based comet assay and 8‐oxodG‐based flow cytometry, the 8‐oxodG expressions were higher in 4βHWE‐treated oral cancer cells than in oral normal cells. All the 4βHWE‐induced DSB and oxidative DNA damage to oral cancer cells were recovered by NAC pretreatment. Taken together, the 4βHWE selectively induced DSB and oxidative DNA damage for the ROS‐mediated selective killing of oral cancer cells.     

Meloxicam inhibits fipronil‐induced apoptosis via modulation of the oxidative stress and inflammatory response in SH‐SY5Y cells

Oxidative stress and inflammatory responses have been identified as key elements of neuronal cell apoptosis. In this study, we investigated the mechanisms by which inflammatory responses contribute to apoptosis in human neuroblastoma SH‐SY5Y cells treated with fipronil (FPN). Based on the cytotoxic mechanism of FPN, we examined the neuroprotective effects of meloxicam against FPN‐induced neuronal cell death. Treatment of SH‐SY5Y cells with FPN induced apoptosis via activation of caspase‐9 and ‐3, leading to nuclear condensation. In addition, FPN induced oxidative stress and increased expression of cyclooxygenase‐2 (COX‐2) and tumor necrosis factor‐α (TNF‐α) via inflammatory stimulation. Pretreatment of cells with meloxicam enhanced the viability of FPN‐exposed cells through attenuation of oxidative stress and inflammatory response. FPN activated mitogen activated protein kinase (MAPK) and inhibitors of MAPK abolished FPN‐induced COX‐2 expression. Meloxicam also attenuated FPN‐induced cell death by reducing MAPK‐mediated pro‐inflammatory factors. Furthermore, we observed both nuclear accumulation of p53 and enhanced levels of cytosolic p53 in a concentration‐dependent manner after FPN treatment. Pretreatment of cells with meloxicam blocked the translocation of p53 from the cytosol to the nucleus. Together, these data suggest that meloxicam may exert anti‐apoptotic effects against FPN‐induced cytotoxicity by both attenuating oxidative stress and inhibiting the inflammatory cascade via inactivation of MAPK and p53 signaling. Copyright © 2015 John Wiley & Sons, Ltd.     

Zinc sulfide nanoparticles selectively induce cytotoxic and genotoxic effects on leukemic cells: involvement of reactive oxygen species and tumor necrosis factor alpha

The aim of the present study was to develop zinc sulfide nanoparticles (ZnS NPs) and to study their cytotoxicity against the KG‐1A (human acute myeloid leukemia) cell line. ZnS NPs were synthesized using the pyrolytic method and characterized by X‐ray diffraction, dynamic light scattering, surface zeta potential, scanning electron microscopy and atomic force microscopy. Cell viability study and flow cytometric analysis confirmed the potent cytotoxic effects of ZnS NPs on cancer cells in a dose‐dependent fashion. Successful uptakes of ZnS NPs by leukemic cells were confirmed by phase contrast fluorescence microscopy. pH‐dependent dissolution of ZnS NPs was done using atomic absorption microscopy to understand the cell‐specific internalization of Zn⁺. This internalization of NPs facilitated the generation of excess reactive oxygen species (ROS), followed by tumor necrosis factor alpha (TNF‐α) secretion which caused severe DNA damage as observed in the comet assay and altered the mitochondrial membrane potential (MMP) in leukemic cells. Surprisingly ZnS NPs had no toxic effects on normal lymphocytes at doses up to 50 µg ml^–1. Pre‐treatment with ROS and TNF‐α inhibitor confirmed that these nanoparticles were able to kill leukemic cells by generating an excess amount of ROS and thereby initiated TNF‐α mediated apoptosis pathway. These findings clarify the mechanism with which ZnS NPs induced anticancer activities in vitro. To elicit its utilities and its application to cancer treatment in vivo is under investigation. Copyright © 2014 John Wiley & Sons, Ltd.     

2, 3, 7, 8‐Tetrachlorodibenzo‐p‐dioxin induces premature senescence of astrocytes via WNT/β‐catenin signaling and ROS production

2, 3, 7, 8‐tetrachlorodibenzo‐p‐dioxin (TCDD) is a ubiquitous environmental contaminant that could exert significant neurotoxicity in the human nervous system. Nevertheless, the molecular mechanism underlying TCDD‐mediated neurotoxicity has not been clarified clearly. Herein, we investigated the potential role of TCDD in facilitating premature senescence in astrocytes and the underlying molecular mechanisms. Using the senescence‐associated β‐galactosidase (SA‐β‐Gal) assay, we demonstrated that TCDD exposure triggered significant premature senescence of astrocyte cells, which was accompanied by a marked activation of the Wingless and int (WNT)/β‐catenin signaling pathway. In addition, TCDD altered the expression of senescence marker proteins, such as p16, p21 and GFAP, which together have been reported to be upregulated in aging astrocytes, in both dose‐ and time‐dependent manners. Further, TCDD led to cell‐cycle arrest, F‐actin reorganization and the accumulation of cellular reactive oxygen species (ROS). Moreover, the ROS scavenger N‐acetylcysteine (NAC) markedly attenuated TCDD‐induced ROS production, cellular oxidative damage and astrocyte senescence. Notably, the application of XAV939, an inhibitor of WNT/β‐catenin signaling pathway, ameliorated the effect of TCDD on cellular β‐catenin level, ROS production, cellular oxidative damage and premature senescence in astrocytes. In summary, our findings indicated that TCDD might induce astrocyte senescence via WNT/β‐catenin and ROS‐dependent mechanisms. Copyright © 2014 John Wiley & Sons, Ltd.     

NF‐κB/p53‐activated inflammatory response involves in diquat‐induced mitochondrial dysfunction and apoptosis

Inflammation generated by environmental toxicants including pesticides could be one of the factors underlying neuronal cell damage in neurodegenerative diseases. In this study, we investigated the mechanisms by which inflammatory responses contribute to apoptosis in PC12 cells treated with diquat. We found that diquat induced apoptosis, as demonstrated by the activation of caspases and nuclear condensation, inhibition of mitochondrial complex I activity, and decreased ATP level in PC12 cells. Diquat also reduced the dopamine level, indicating that cell death induced by diquat is due to cytotoxicity of dopaminergic neuronal components in these cells. Exposure of PC12 cells to diquat led to the production of reactive oxygen species (ROS), and the antioxidant N‐acetyl‐cystein attenuated the cytotoxicity of caspase‐3 pathways. These results demonstrate that diquat‐induced apoptosis is involved in mitochondrial dysfunction through production of ROS. Furthermore, diquat increased expression of cyclooxygenase‐2 (COX‐2) and tumor necrosis factor‐α (TNF‐α) via inflammatory stimulation. Diquat induced nuclear accumulation of NF‐κB and p53 proteins. Importantly, an inhibitor of NF‐κB nuclear translocation blocked the increase of p53. Both NF‐κB and p53 inhibitors also blocked the diquat‐induced inflammatory response. Pretreatment of cells with meloxicam, a COX‐2 inhibitor, also blocked apoptosis and mitochondrial dysfunction. These results represent a unique molecular characterization of diquat‐induced cytotoxicity in PC12 cells. Our results demonstrate that diquat induces cell damage in part through inflammatory responses via NF‐κB‐mediated p53 signaling. This suggests the potential to generate mitochondrial damage via inflammatory responses and inflammatory stimulation‐related neurodegenerative disease.     

Galectin‐3 aggravates ox‐LDL‐induced endothelial dysfunction through LOX‐1 mediated signaling pathway

Galectin‐3, a biomarker linking oxidative stress and inflammation, participates in different mechanisms related to atherothrombosis, such as inflammation, proliferation, or macrophage chemotaxis. Accumulating evidence indicates that galectin‐3 may also promote atherogenesis through inducing endothelial dysfunction. Lectin‐like oxidized low‐density lipoprotein (oxLDL) receptor‐1 (LOX‐1), a receptor for oxLDL uptake, contributes to oxLDL‐induced endothelial dysfunction. Whether galectin‐3 induces endothelial dysfunction through modulation of LOX‐1‐mediated signaling remains unclear. In the present study, we explored the mechanisms underlying galectin‐3 enhanced cytotoxicity of oxLDL in human umbilical vein endothelial cells (HUVECs) and the role of LOX‐1. Incubation of HUVECs with galectin‐3 increased the expression of LOX‐1 in RNA and protein levels. In addition, the expression of LOX‐1 induced by oxLDL was promoted by galectin‐3. However, pretreatment of LOX‐1 antibody reduced LOX‐1 mRNA expression level in cells with oxLDL plus galectin‐3 incubation. Compared to cells treated with oxLDL alone, reactive oxygen species (ROS) generation via nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation and subsequent activation of p38 mitogen‐activated protein kinases followed by nuclear factor kappa B (NF‐κB) activation and related inflammatory responses including adhesion molecule expression, adhesiveness of monocytic cells, and IL‐8 release were also aggravated in cells treated with galectin‐3 combined with oxLDL. Compared to cells treated with galectin‐3 plus oxLDL group. We found that LOX‐1 antibody mitigated NADPH oxidase activity, p‐38 up‐regulation, NF‐κB activation, and proinflammatory responses in cells treated with galectin‐3 combined with oxLDL. We conclude that galectin‐3 enhances endothelial LOX‐1 expression and propose a new mechanism by which galectin‐3 may promote endothelial dysfunction by inducing inflammation via LOX‐1/ROS/p38/NF‐κB‐mediated signaling pathway.     

Cantharidic acid induces apoptosis of human leukemic HL‐60 cells via c‐Jun N‐terminal kinase‐regulated caspase‐8/‐9/‐3 activation pathway

Cantharidin, a natural toxin from blister beetles, has shown potent anticancer activities on many solid tumor cells. Recently, cantharidin and its analogue, norcantharidin, were also shown to suppress nonsolid tumors such as chronic myeloid leukemia, acute myeloid leukemia (AML), and leukemic stem cells. However, there is no available information to address the effects of cantharidic acid (CAC), a hydrolysis product of cantharidin, on human AML cells. The present study showed that CAC, at a range of concentrations (0‐20 μM), concentration‐dependently inhibited cell proliferation in the HL‐60 AML cell line. Western blot and flow cytometric assays demonstrated that CAC induced several features of apoptosis such as sub G1‐phase cell increase, phosphatidylserine (PS) externalization, and significantly activated proapoptotic signaling including caspase‐8, −9, and −3 activation and poly(ADP‐ribose) polymerase (PARP) cleavage in HL‐60 AML cells. Moreover, treatment of HL‐60 cells with CAC induced concentration‐ and time‐ dependent activation of p38 mitogen‐activated protein kinase (p38 MAPK) and c‐Jun N‐terminal kinase (JNK). Only JNK‐, but not p38 MAPK‐specific inhibitor can reverse the CAC‐induced activation of the caspase‐8, −9, and −3. We concluded that CAC can induce apoptosis in human leukemic HL‐60 cells via a caspases‐dependent pathway, and that the apoptosis‐inducing effect of CAC can be regulated by JNK activation signaling.     

NOX4‐ and Nrf2‐mediated oxidative stress induced by silver nanoparticles in vascular endothelial cells

It has been widely reported that silver nanoparticles (AgNPs) induce oxidative stress in various cell lines. However, the mechanism for this effect and its consequences for cellular signaling are poorly understood. In this study, human umbilical vein endothelial cells (HUVECs) were used to assess the toxicity and investigate the associated molecular mechanisms caused by exposure to AgNPs. We demonstrated that AgNP exposure significantly and dose‐dependently decreased the cell viability, induced reactive oxygen species (ROS) generation and led to early apoptosis in HUVECs. Our findings showed that AgNPs induced excess ROS production that affected the signaling pathways by a mechanism that depended on activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity through upregulation of NADPH oxidase 4 (NOX4) protein expressions. Moreover, AgNPs could disrupt the inactivation of the nuclear factor erythroid 2‐related factor 2 (Nrf2)‐mediated antioxidant response, which is considered another important element for oxidative stress caused by AgNPs in HUVECs. The redox imbalance between NOX4 and Nrf2 was an important cause for the ROS overproduction that led to cell injury in HUVECs. The results provided insight into the mechanisms of oxidative stress induced by AgNPs in vascular endothelial cells.     

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin (TCDD)‐Induced Activation of Mitogen‐Activated Protein Kinase Signaling Pathway in Jurkat T Cells

Abstract: The present study was performed to examine mitogen‐activated protein kinase associated pathways in mediation of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD)‐induced cell apoptosis in cultured Jurkat T cells. TCDD significantly decreased cell viability in a concentration‐dependent manner (P<0.05 at 10–300 nM). TCDD (10 nM) also time‐dependently decreased cell viability (P<0.05 at 12–48 hr). c‐Jun NH₂‐terminal kinase was significantly phosphorylated with TCDD treatment in a time dependent manner. p38 Mitogen‐activated protein kinase was not significantly changed with TCDD treatment. Extracellular signal‐regulated protein kinase was significantly phosphorylated with TCDD treatment for 8 hr and gradually returned to baseline. TCDD induced up‐regulation of ASK1 and C‐Jun, which are up‐ and down‐stream of JNK, respectively, and up‐regulation of cytosolic cytochrome c and caspase‐3. These results demonstrate that MAPK signaling pathways including JNK and ERK 1/2, are activated with the treatment of TCDD in Jurkat T cells, which suggest that MAPK pathways may be involved in TCDD‐induced cell death.     

Fisetin‐induced apoptosis of human oral cancer SCC‐4 cells through reactive oxygen species production,endoplasmic reticulum stress,caspase‐, and mitochondria‐dependent signaling pathways

Oral cancer is one of the cancer‐related diseases in human populations and its incidence rates are rising worldwide. Fisetin, a flavonoid from natural products, has been shown to exhibit anticancer activities in many human cancer cell lines but the molecular mechanism of fisetin‐induced apoptosis in human oral cancer cells is still unclear; thus, in this study, we investigated fisetin‐induced cell death and associated signal pathways on human oral cancer SCC‐4 cells in vitro. We examined cell morphological changes, total viable cells, and cell cycle distribution by phase contrast microscopy and flow cytometry assays. Reactive oxygen species (ROS), Ca²⁺, mitochondria membrane potential (ΔΨ_m), and caspase‐8, ‐9, and ‐3 activities were also measured by flow cytometer. Results indicate that fisetin induced cell death through the cell morphological changes, caused G2/M phase arrest, induction of apoptosis, promoted ROS and Ca²⁺ production, and decreased the level of ΔΨ_m and increased caspase‐3, ‐8, and ‐9 activities in SCC‐4 cells. DAPI staining and DNA gel electrophoresis were also used to confirm fisetin‐induced cell apoptosis in SCC‐4 cells. Western blotting also found out that Fisetin increased the proapoptotic proteins such as Bax and Bid and decreased the antiapoptotic proteins such as Bcl‐2. Furthermore, results also showed that Fisetin increased the cytochrome c, AIF, and Endo G release from mitochondria in SCC‐4 cells. We also used ATF‐6α, ATF‐6β, GADD153, and GRP78 which indicated that fisetin induced cell death through ER stress. Based on those observations, we suggest that fisetin induced cell apoptosis through ER stress, mitochondria‐, and caspase‐dependent pathways.     

Microcystin‐LR‐Caused ROS generation involved in p38 activation and tau hyperphosphorylation in neuroendocrine (PC12) cells

Microcystin‐LR (MC‐LR), a potent specific hepatotoxin produced by cyanobacteria, has recently been reported to show neurotoxicity. Our previous study demonstrated that MC‐LR caused the reorganization of cytoskeleton architectures and hyperphosphorylation of the cytoskeletal‐associated proteins tau and HSP27 in neuroendocrine PC12 cell line by direct PP2A inhibition and indirect p38 mitogen‐activated protein kinase (MAPK) activation. It has been shown that oxidative stress is extensively associated with MC‐LR toxicity, mainly resulting from an excessive production of reactive oxygen species (ROS). However, the mechanisms by which ROS mediates the cytotoxic action of MC‐LR are unclear. In the present study, we investigated whether ROS might play a critical role in MC‐LR‐induced hyperphosphorylation of microtubule‐associated protein tau and the activation of the MAPKs in PC12 cell line. The results showed that MC‐LR had time‐ and concentration‐dependent effects on ROS generation, p38‐MAPK activation and tau phosphorylation. The time‐course studies indicated similar biphasic changes in ROS generation and tau hyperphosphorylation, which started to increase within 1 h and reached the maximum level at 3 h followed by a decrease after prolonged treatment. Furthermore, pretreatment with the antioxidants, N‐acetylcysteine and vitamin C, significantly decreased MC‐LR‐induced ROS generation and effectively attenuated p38‐MAPK activation as well as tau hyperphosphorylation. Taken together, these findings suggest that ROS generation triggered by MC‐LR is a key intracellular event that contributes to an induction of p38‐MAPK activation and tau phosphorylation, and that blockade of this ROS‐mediated redox‐sensitive signal cascades may attenuate the toxic effects of MC‐LR. © 2013 Wiley Periodicals, Inc. Environ Toxicol 30: 366–374, 2015.     

Mechanistic insight into reactivity and (geno)toxicity of well-characterized nanoparticles of cobalt metal and oxides

An increasing use of cobalt (Co)-based nanoparticles (NPs) in different applications and exposures at occupational settings triggers the need for toxicity assessment. Improved understanding regarding the physiochemical characteristics of Co metal NPs and different oxides in combination with assessment of toxicity and mechanisms may facilitate decisions for grouping during risk assessment. The aim of this study was to gain mechanistic insights in the correlation between NP reactivity and toxicity of three different Co-based NPs (Co, CoO, and Co₃O₄) by using various tools for characterization, traditional toxicity assays, as well as six reporter cell lines (ToxTracker) for rapid detection of signaling pathways of relevance for carcinogenicity. The results showed cellular uptake of all NPs in lung cells and induction of DNA strand breaks and oxidative damage (comet assay) by Co and CoO NPs. In-depth studies on the ROS generation showed high reactivity of Co, lower for CoO, and no reactivity of Co₃O₄ NPs. The reactivity depended on the corrosion and transformation/dissolution properties of the particles and the media highlighting the role of the surface oxide and metal speciation as also confirmed by in silico modeling. By using ToxTracker, Co NPs were shown to be highly cytotoxic and induced reporters related to oxidative stress (Nrf2 signaling) and DNA strand breaks. Similar effects were observed for CoO NPs but at higher concentrations, whereas the Co₃O₄ NPs were inactive at all concentrations tested. In conclusion, our study suggests that Co and CoO NPs, but not Co₃O₄, may be grouped together for risk assessment.     

The role of oxidative stress in citreoviridin‐induced DNA damage in human liver‐derived HepG2 cells

We hypothesize that citreoviridin (CIT) induces DNA damage in human liver‐derived HepG2 cells through an oxidative stress mechanism and that N‐acetyl‐l ‐cysteine (NAC) protects against CIT‐induced DNA damage in HepG2 cells. CIT‐induced DNA damage in HepG2 cells was evaluated by alkaline single‐cell gel electrophoresis assay. To elucidate the genotoxicity mechanisms, the level of oxidative DNA damage was tested by immunoperoxidase staining for 8‐hydroxydeoxyguanosine (8‐OHdG); the intracellular generation of reactive oxygen species (ROS) and reduced glutathione (GSH) were examined; mitochondrial membrane potential and lysosomal membranes' permeability were detected; furthermore, protective effects of NAC on CIT‐induced ROS formation and CIT‐induced DNA damage were evaluated in HepG2 cells. A significant dose‐dependent increment in DNA migration was observed at tested concentrations (2.50–10.00 µM) of CIT. The levels of ROS, 8‐OHdG formation were increased by CIT, and significant depletion of GSH in HepG2 cells was induced by CIT. Destabilization of lysosome and mitochondria was also observed in cells treated with CIT. In addition, NAC significantly decreased CIT‐induced ROS formation and CIT‐induced DNA damage in HepG2 cells. The data indicate that CIT induces DNA damage in HepG2 cells, most likely through oxidative stress mechanisms; that NAC protects against DNA damage induced by CIT in HepG2 cells; and that depolarization of mitochondria and lysosomal protease leakage may play a role in CIT‐induced DNA damage in HepG2 cells. © 2014 The Authors. Published by Wiley Periodicals Inc. Environ Toxicol 30: 530–537, 2015.     

Copper nanoparticle‐induced uterine injury in female rats

Copper nanoparticles (Cu‐NPs) have been used increasingly in various products and applications. Although recent studies have reported that exposure to Cu‐NPs leads to organ accumulation and obvious toxicity, it remains unclear whether Cu‐NPs can be translocated to and cause damage in the uterus. In this study, we investigated the potential for uterine injury and gene expression patterns in female rats exposed to 3.12, 6.25, or 12.5 mg/kg/d Cu‐NPs via intraperitoneal injection for 14 consecutive days. The results indicated that exposure to Cu‐NPs led to significant decreases in the relative uterine weight coefficients and increases in inflammatory cell infiltration, mitochondrial swelling and vacuolization, shortened and reduced endometrial epithelial cell microvilli, and apoptosis. Furthermore, exposure to Cu‐NPs increased malondialdehyde (MDA) accumulation and decreased superoxide dismutase (SOD) levels. Signal transduction mechanism studies indicated that exposure to Cu‐NPs activated caspases 3, 8, and 9 and BH3 interacting domain death agonist (tBid), reduced B cell leukemia/lymphoma 2 (Bcl‐2) expression, and increased the expression of apoptotic peptidase activating factor 1 (Apaf‐1), BCL2‐associated X, apoptosis regulator (Bax), and cytochrome c. A microarray analysis revealed significant alterations in the expression of 963 genes; of these, 622 were upregulated and 341 were downregulated. The results of further evaluations of some altered genes, including matrix metallopeptidase 12 (Mmp12), using quantitative RT‐PCR agreed with the microarray findings. These results provide strong evidence that Cu‐NPs can trigger both intrinsic and extrinsic apoptotic pathways to mediate uterine injury, resulting in oxidative stress‐related changes in gene expression.     

Mono(2‐ethylhexyl) phthalate induces apoptosis in p53‐silenced L02 cells via activation of both mitochondrial and death receptor pathways

Mono(2‐ethylhexyl) phthalate (MEHP) is one of the main metabolites of di(2‐ethylhexyl) phthalate. The evidence shows that DEHP may exert its toxic effects primarily via MEHP, which is 10‐fold more potent than its parent compound in toxicity in vitro. MEHP‐induced apoptosis is mediated by either p53‐dependent or ‐independent pathway. However, the detailed mechanism of its toxicity remains unclear. In this study, immortalized normal human liver cell line L02 was chosen, as an in vitro model of nonmalignant liver, to elucidate the role of p53 in MEHP‐induced apoptosis. The cells were treated with MEHP (6.25, 12.50, 25.00, 50.00, and 100.00 μM) for 24 and 36 h, then small interfering RNA (siRNA) was used to specifically silence p53 gene of L02 cells. The results indicated that MEHP caused oxidative DNA damage and apoptosis in L02 cells were associated with the p53 signaling pathway. Further study found that MEHP (50.00 and 100.00 μM) induced apoptosis in p53‐silenced L02 cells, along with the up‐regulations of Fas and FasL proteins as well as increased the Bax/Bcl‐2 ratio and Caspase 3, 8, and 9 activities. Additionally, both FasL inhibitor (AF‐016) and Caspase inhibitor N‐benzyloxycarbonyl‐Val‐Ala‐Asp‐ fluoromethylketone (Z‐VAD‐FMK) could prevent the cell apoptosis induced by MEHP. The findings suggested that MEHP‐induced apoptosis in L02 cells involving a Caspases‐mediated mitochondrial signaling pathway and/or death receptor pathway. p53 was not absolutely necessary for MEHP‐induced L02 cell apoptosis. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1178–1191, 2015.     

3‐Hydroxyflavone enhances the toxicity of ZnO nanoparticles in vitro

It is recently shown that flavonoids might reduce the toxicity of nanoparticles (NPs) due to their antioxidative properties. In this study, the influence of 3‐hydroxyflavone (H3) on the toxicity of ZnO NPs was investigated. H3 increased hydrodynamic size, polydispersity index and absolute value of the zeta potential of ZnO NPs, which indicated that H3 could influence the colloidal aspects of NPs. Surprisingly, H3 markedly decreased the initial concentration of ZnO NPs required to induce cytotoxicity to Caco‐2, HepG2, THP‐1 and human umbilical vein endothelial cells, which suggested that H3 could promote the toxicity of ZnO NPs to both cancerous and normal cells. For comparison, 6‐hydroxyflavone did not show this effect. H3 remarkably increased cellular Zn elements and intracellular Zn ions in HepG2 cells following ZnO NP exposure, and co‐exposure to H3 and NPs induced a relatively higher intracellular reactive oxygen species. Exposure to ZnO NPs at 3 hours induced the expression of endoplasmic reticulum stress markers DDIT3 and XBP‐1 s, which was suppressed by H3. The expression of apoptotic genes BAX and CASP3 was significantly induced by ZnO NP exposure after 3 and 5 hours, respectively, and H3 further significantly promoted CASP3 expression at 5 hours. In combination, the results from this study suggested that H3 affected colloidal stability of ZnO NPs, promoted the interactions between NPs and cells, and altered the NP‐induced endoplasmic reticulum stress–apoptosis signaling pathway, which finally enhanced the cytotoxicity of ZnO NPs.     

3‐Keto‐1,5‐bisphosphonates Alleviate Serum‐Oxidative Stress in the High‐fat Diet Induced Obesity in Rats

Obesity has become a leading global health problem owing to its strong association with a high incidence of oxidative stress. Many epidemiologic studies showed that an antioxidant supplementation decreases the state of oxidative stress. In the present work, a HFD ‐induced rat obesity and oxidative stress were used to investigate the link between fat deposition and serum‐oxidative stress markers. We also studied the effect of a chronic administration of 3‐keto‐1,5‐bisphosphonates 1 (a & b) (40 μg/kg/8 weeks/i.p.). Exposure of rats to HFD during 16 weeks induced fat deposition, weight gain and metabolic disruption characterized by an increase in cholesterol, triglyceride and glycemia levels, and a decrease in ionizable calcium and free iron concentrations. HFD also induced serum‐oxidative stress status vocalized by an increase in ROS ( H ₂ O ₂ ), MDA and PC levels, with a decrease in antioxidant enzyme activity ( CAT , GP x , SOD ). Importantly, 3‐keto‐1,5‐bisphosphonates corrected all the deleterious effects of HFD treatment in vivo, but it failed to inhibit lipases in vitro and in vivo. These studies suggest that 3‐keto‐1,5‐bisphosphonates 1 could be considered as safe antioxidant agents that should also find other potential biological applications.