The fumonisins are mycotoxins produced by fungi that contaminate primarily corn and are toxic through interruption of intracellular sphingolipid synthesis. Several reports have indicated that fumonisin B1 concentrations decreased when heated in aqueous solutions of reducing sugars. The incubation of fumonisin B1 with d-glucose resulted in the formation of N-(carboxymethyl)fumonisin B1, which was characterized by NMR and electrospray mass spectroscopy. We determined the methylene carbon of the carboxymethyl group is derived from C1 on glucose, while the carbonyl carbon is derived from the C2 of glucose, using 13C glucose. Apparently N-(carboxymethyl)fumonisin B1 arises from Schiff's base formation, Amadori rearrangement to a β-ketoamine, and oxidation with molecular oxygen. N-(Carboxymethyl)fumonisin B1 formation is favored by alkaline conditions (pH >7), requires molecular oxygen, and is catalyzed by several reducing sugars. N-(carboxymethyl)fumonisin B1 was detected in raw corn samples that contained fumonisin B1 (0.5−1.4 ppm) at an average of 4% of the fumonisin B1 levels. Keywords: Fumonisin B1; fumonisins; N-(carboxymethyl)fumonisin B1; detoxification; Amadori rearrangement; Maillard reaction; reducing sugars; mycotoxin

Sertraline, a selective serotonin reuptake inhibitor, has been used for the treatment of depression. Although it is generally considered safe, cases of sertraline-associated liver injury have been documented; however, the possible mechanism of sertraline-associated hepatotoxicity is entirely unknown. Here, we report that mitochondrial impairment may play an important role in liver injury induced by sertraline. In mitochondria isolated from rat liver, sertraline uncoupled mitochondrial oxidative phosphorylation and inhibited the activities of oxidative phosphorylation complexes I and V. Additionally, sertraline induced Ca(2+)-mediated mitochondrial permeability transition (MPT), and the induction was prevented by bongkrekic acid (BA), a specific MPT inhibitor targeting adenine nucleotide translocator (ANT), implying that the MPT induction is mediated by ANT. In freshly isolated rat primary hepatocytes, sertraline rapidly depleted cellular adenosine triphosphate (ATP) and subsequently induced lactate dehydrogenase leakage; both were attenuated by BA. Our results, including ATP depletion, induction of MPT, inhibition of mitochondrial respiration complexes, and uncoupling oxidative phosphorylation, indicate that sertraline-associated liver toxicity is possibly via mitochondrial dysfunction.

Sertraline is generally used for the treatment of depression and is also approved for the treatment of panic, obsessive-compulsive, and posttraumatic stress disorders. Previously, using rat primary hepatocytes and isolated mitochondria, we demonstrated that sertraline caused hepatic cytotoxicity and mitochondrial impairment. In the current study, we investigated and characterized molecular mechanisms of sertraline toxicity in human hepatoma HepG2 cells. Sertraline decreased cell viability and induced apoptosis in a dose- and time-dependent manner. Sertraline activated the intrinsic checkpoint protein caspase-9 and caused the release of cytochrome c from mitochondria to cytosol; this process was Bcl-2 family dependent because antiapoptotic Bcl-2 family proteins were decreased. Pretreatment of the HepG2 cells with caspase-3, caspase-8, and caspase-9 inhibitors partially but significantly reduced the release of lactate dehydrogenase, indicating that sertraline-induced apoptosis is mediated by both intrinsic and extrinsic apoptotic pathways. Moreover, sertraline markedly increased the expression of tumor necrosis factor (TNF) and the phosphorylation of JNK, extracellular signal-regulated kinase (ERK1/2), and p38. In sertraline-treated cells, the induction of apoptosis and cell death was shown to be the result of activation of JNK, but not ERK1/2 or p38 in the mitogen-activated protein kinase (MAPK) pathway. Furthermore, silencing MAP4K4, the upstream kinase of JNK, attenuated both apoptosis and cell death caused by sertraline. Taken together, our findings suggest that sertraline induced apoptosis in HepG2 cells at least partially via activation of the TNF-MAP4K4-JNK cascade signaling pathway.

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SUMMARY The role of mucosal fibroblasts in intestinal inflammatory reactions is not known. In this study, we demonstrate that fibroblasts grown from histologically normal human duodenal biopsy tissues expressed mRNA genes for granulocyte‐macrophage colony‐stimulating factor (GM‐CSF). IL‐lα, IL‐1β, IL‐6, IL‐8, IL‐10, intercellular adhesion molecule‐I (ICAM‐I) and vascular cell adhesion molecule‐1 (VCAM‐i) when stimulated with lipopolysaccharide (LPS) or IL‐1α. The increased mRNA expression of GM‐CSF, IL‐1α IL‐1β IL‐6 and IL‐α in response to IL‐1α and LPS stimulation was time‐ and dose‐dependent. In contrast. IL‐10 was weakly expressed when fibroblasts were stimulated with LPS. IL‐1α or tumour necrosis factor‐alpha (TNF‐α), but the expression was enhanced in the presence of cycloheximide combined with optimal concentrations of LPS. IL‐1α or TNF‐α. IL‐1α was a more potent stimulator than LPS for GM‐CSF. IL‐6, IL‐8 and I L‐10 expression, but not for IL‐1α and IL‐1β. Increased GM‐CSF. lL‐6 and IL‐8 gene expression was associated with the production of cytokine proteins in culture supernatant, but IL‐1α and IL‐1bL remained undetectable. Dexamethasone suppressed both gene expression and protein production of GM‐CSF. IL‐6 and IL‐8 when fibroblasts were exposed to IL‐1α. TNF‐α stimulated the release of GM‐CSF. IL‐6 and IL‐8 and, combined with IL‐1α. cytokine production was enhanced synergistically. Finally, both LPS and IL‐1ã up‐regulated ICAM‐I and VCAM‐1 gene expression. These findings implicate duodenal fibroblasts in the initiation and/or regulation of intestinal inflammation.

Ginkgo biloba leaf extract has been shown to increase the incidence in liver tumors in mice in a 2-year bioassay conducted by the National Toxicology Program. In this study, the DNA damaging effects of Ginkgo biloba leaf extract and many of its constituents were evaluated in human hepatic HepG2 cells and the underlying mechanism was determined. A molecular docking study revealed that quercetin, a flavonoid constituent of Ginkgo biloba, showed a higher potential to interact with topoisomerase II (Topo II) than did the other Ginkgo biloba constituents; this in silico prediction was confirmed by using a biochemical assay to study Topo II enzyme inhibition. Moreover, as measured by the Comet assay and the induction of γ-H2A.X, quercetin, followed by keampferol and isorhamnetin, appeared to be the most potent DNA damage inducer in HepG2 cells. In Topo II knockdown cells, DNA damage triggered by Ginkgo biloba leaf extract or quercetin was dramatically decreased, indicating that DNA damage is directly associated with Topo II. DNA damage was also observed when cells were treated with commercially available Ginkgo biloba extract product. Our findings suggest that Ginkgo biloba leaf extract- and quercetin-induced in vitro genotoxicity may be the result of Topo II inhibition.

Sertraline is used for the treatment of depression, and is also used for the treatment of panic, obsessive-compulsive, and post-traumatic stress disorders. Previously, we have demonstrated that sertraline caused hepatic cytotoxicity, with mitochondrial dysfunction and apoptosis being underlying mechanisms. In this study, we used microarray and other biochemical and molecular analyses to identify endoplasmic reticulum (ER) stress as a novel molecular mechanism. HepG2 cells were exposed to sertraline and subjected to whole genome gene expression microarray analysis. Pathway analysis revealed that ER stress is among the significantly affected biological changes. We confirmed the increased expression of ER stress makers by real-time PCR and Western blots. The expression of typical ER stress markers such as PERK, IRE1α, and CHOP was significantly increased. To study better ER stress-mediated drug-induced liver toxicity; we established in vitro systems for monitoring ER stress quantitatively and efficiently, using Gaussia luciferase (Gluc) and secreted alkaline phosphatase (SEAP) as ER stress reporters. These in vitro systems were validated using well-known ER stress inducers. In these two reporter assays, sertraline inhibited the secretion of Gluc and SEAP. Moreover, we demonstrated that sertraline-induced apoptosis was coupled to ER stress and that the apoptotic effect was attenuated by 4-phenylbutyrate, a potent ER stress inhibitor. In addition, we showed that the MAP4K4-JNK signaling pathway contributed to the process of sertraline-induced ER stress. In summary, we demonstrated that ER stress is a mechanism of sertraline-induced liver toxicity.

Tattooing has become a popular recreational practice among younger adults over the past decade. Although some of the pigments used in tattooing have been described, very little is known concerning the toxicology, phototoxicology or photochemistry of these pigments. Seven yellow tattoo inks were obtained from commercial sources and their pigments extracted, identified and quantitatively analyzed. The monoazo compound Pigment Yellow 74 (PY74; CI 11741) was found to be the major pigment in several of the tattoo inks. Solutions of commercial PY74 in tetrahydrofuran (THF) were deoxygenated using argon gas, and the photochemical reaction products were determined after exposure to simulated solar light generated by a filtered 6.5 kW xenon arc lamp. Spectrophotometric and high-pressure liquid chromatography (HPLC) analyses indicated that PY74 photodecomposed to multiple products that were isolated using a combination of silica chromatography and reversed-phase HPLC. Three of the major photodecomposition products were identified by nuclear magnetic resonance and mass spectrometry as N-(2-methoxyphenyl)-3-oxobutanamide (o-acetoacetanisidide), 2-(hydroxyimine)-N-(2-methoxyphenyl)-3-oxobutanamide and N,N''-bis(2-methoxyphenyl)urea. These results demonstrate that PY74 is not photostable in THF and that photochemical lysis occurs at several sites in PY74 including the hydrazone and amide groups. The data also suggest that the use of PY74 in tattoo inks could potentially result in the formation of photolysis products, resulting in toxicity at the tattoo site after irradiation with sunlight or more intense light sources.