Metalloprotein-Specific or Critical Amino Acid Residues: Perspectives on Plant-Precise Detoxification and Recognition Mechanisms under Cadmium Stress.

Cadmium (Cd) pollution in cultivated land is caused by irresistible geological factors and human activities; intense diffusion and migration have seriously affected the safety of food crops. Plants have evolved mechanisms to control excessive influx of Cd in the environment, such as directional transport, chelation and detoxification. This is done by some specific metalloproteins, whose key amino acid motifs have been investigated by scientists one by one. The application of powerful cell biology, crystal structure science, and molecular probe targeted labeling technology has identified a series of protein families involved in the influx, transport and detoxification of the heavy metal Cd. This review summarizes them as influx proteins (NRAMP, ZIP), chelating proteins (MT, PDF), vacuolar proteins (CAX, ABCC, MTP), long-distance transport proteins (OPT, HMA) and efflux proteins (PCR, ABCG). We selected representative proteins from each family, and compared their amino acid sequence, motif structure, subcellular location, tissue specific distribution and other characteristics of differences and common points, so as to summarize the key residues of the Cd binding target. Then, we explain its special mechanism of action from the molecular structure. In conclusion, this review is expected to provide a reference for the exploration of key amino acid targets of Cd, and lay a foundation for the intelligent design and breeding of crops with high/low Cd accumulation.

CYP2C9 and 3A4 play opposing roles in bioactivation and detoxification of diphenylamine NSAIDs.

Diphenylamine NSAIDs are taken frequently for chronic pain conditions, yet their use may potentiate hepatotoxicity risks through poorly characterized metabolic mechanisms. Our previous work revealed that seven marketed or withdrawn diphenylamine NSAIDs undergo bioactivation into quinone-species metabolites, whose reaction specificities depended on halogenation and the type of acidic group on the diphenylamine. Herein, we identified cytochromes P450 responsible for those bioactivations, determined reaction specificities, and estimated relative contributions of enzymes to overall hepatic bioactivations and detoxifications. A qualitative activity screen revealed CYP2C8, 2C9, 2C19, and 3A4 played roles in drug bioactivation. Subsequent steady-state studies with recombinant CYPs recapitulated the importance of halogenation and acidic group type on bioactivations but importantly, showed patterns unique to each CYP. CYP2C9, 2C19 and 3A4 bioactivated all NSAIDs with CYP2C9 dominating all possible bioactivation pathways. For each CYP, specificities for overall oxidative metabolism were not impacted significantly by differences in NSAID structures but the values themselves differed among the enzymes such that CYP2C9 and 3A4 were more efficient than others. When considering hepatic CYP abundance, CYP2C9 almost exclusively accounted for diphenylamine NSAID bioactivations, whereas CYP3A4 provided a critical counterbalance favoring their overall detoxification. Preference for either outcome would depend on molecular structures favoring metabolism by the CYPs as well as the influence of clinical factors altering their expression and/or activity. While focused on NSAIDs, these findings have broader implications on bioactivation risks given the expansion of the diphenylamine scaffold to other drug classes such as targeted cancer therapeutics.

Toxicity of Bioactive Molecule Andrographolide against Spodoptera litura Fab and Its Binding Potential with Detoxifying Enzyme Cytochrome P450.

Spodoptera litura Fab. is a polyphagous pest causing damage to many agriculture crops leading to yield loss. Recurrent usage of synthetic pesticides to control this pest has resulted in resistance development. Plant-derived diterpenoid compound andrographolide was isolated from the leaves of Andrographis paniculata. It was analysed by gas chromatography-mass spectroscopy and quantified by HPLC. Nutritional indices and digestive enzymatic profile were evaluated. Third, fourth and fifth instar larvae were treated with different concentrations of andrographolide. At 3, 6 and 9 ppm-treated concentrations the larvae showed decreased RGR, RCR, ECI, ECD values with adverse increase in AD. The digestive enzymes were significantly inhibited when compared with control. Conspicuously, andrographolide showed pronounced mortality of S. litura by inhibition of enzyme secretion and intake of food. The binding ability of andrographolide with CYTP450 showed high affinity with low binding energy. Andrographolide has the potential to be exploited as a biocontrol agent against S. litura as an eco-friendly pesticide.

Quantitative Investigation of Irinotecan Metabolism, Transport, and Gut Microbiome Activation.

The anticancer drug irinotecan shows serious dose-limiting gastrointestinal toxicity regardless of intravenous dosing. Although enzymes and transporters involved in irinotecan disposition are known, quantitative contributions of these mechanisms in complex in vivo disposition of irinotecan are poorly understood. We explained intestinal disposition and toxicity of irinotecan by integrating 1) in vitro metabolism and transport data of irinotecan and its metabolites, 2) ex vivo gut microbial activation of the toxic metabolite SN-38, and 3) the tissue protein abundance data of enzymes and transporters relevant to irinotecan and its metabolites. Integration of in vitro kinetics data with the tissue enzyme and transporter abundance predicted that carboxylesterase (CES)-mediated hydrolysis of irinotecan is the rate-limiting process in the liver, where the toxic metabolite formed is rapidly deactivated by glucuronidation. In contrast, the poor SN-38 glucuronidation rate as compared with its efficient formation by CES2 in the enterocytes is the key mechanism of the intestinal accumulation of the toxic metabolite. The biliary efflux and organic anion transporting polypeptide-2B1-mediated enterocyte uptake can also synergize buildup of SN-38 in the enterocytes, whereas intestinal P-glycoprotein likely facilitates SN-38 detoxification in the enterocytes. The higher SN-38 concentration in the intestine can be further nourished by ß-d-glucuronidases. Understanding the quantitative significance of the key metabolism and transport processes of irinotecan and its metabolites can be leveraged to alleviate its intestinal side effects. Further, the proteomics-informed quantitative approach to determine intracellular disposition can be extended to determine susceptibility of cancer cells over normal cells for precision irinotecan therapy. SIGNIFICANCE STATEMENT: This work provides a deeper insight into the quantitative relevance of irinotecan hydrolysis (activation), conjugation (deactivation), and deconjugation (reactivation) by human or gut microbial enzymes or transporters. The results of this study explain the characteristic intestinal exposure and toxicity of irinotecan. The quantitative tissue-specific in vitro to in vivo extrapolation approach presented in this study can be extended to cancer cells.

Detoxification of Oral Exposure to Benzo(a)pyrene by Lactobacillus plantarum CICC 23121 in Mice.

This study's previous work showed that the carcinogen and mutagen benzo(a)pyrene (BaP) can be adsorbed by Lactobacillus cells in vitro. However, in vivo BaP detoxification by lactic acid bacteria has not yet been investigated. The present study evaluates the effects of orally administered Lactobacillus plantarum CICC 23121 in BaP-treated mice. Oral administration of 50 mg kg-1 BaP perturbed the intestinal microflora, caused Proteobacteria to predominate, and severely damaged DNA. However, oral administration of 5 × 1010 CFU mL-1 CICC 23121 in BaP-treated mice enhances fecal BaP excretion from 181.70 ± 1.04 µg/(g∙h) to 271.47 ± 11.71 µg/(g∙h) after 6 h. Fecal BaP excretion reaches up to 280.66 ± 22.97 µg/(g∙h) after the first 4 days of orally administered CICC 23121 and decreased to 94.31 ± 2.64 µg/(g∙h) by day 11. Intestinal microbiota are restored and Firmicutes predominates. CICC 23121 alleviates BaP-induced DNA damage and reduces tail length from 56.37 ± 5.31  to 39.69 ± 4.27 µm. Therefore, oral CICC23121 consumption is a promising strategy for reducing BaP toxicity in mice. To the best of our knowledge, this report is the first report to demonstrate in vivo that Lactobacillus cells can detoxify BaP.

Melatonin alleviates imidacloprid phytotoxicity to cucumber (Cucumis sativus L.) through modulating redox homeostasis in plants and promoting its metabolism by enhancing glutathione dependent detoxification.

Melatonin (Mel), a powerful antioxidant that has the ability to regulate physiological and biochemical processes in plants under abiotic stresses. However, its roles in pesticide detoxification is poorly understood. Herein, selecting leaf spraying insecticide imidacloprid (IMD) as the model, we demonstrated the detoxification mechanism underlying root pretreatment of Mel on IMD in cucumber. IMD treatment affected the primary light conversion efficiency of photosystem II (Fv/Fm), reduced the quantum yield, and increased hydrogen peroxide and superoxide anions contents as well as the levels of membrane lipid peroxidation, indicating that excessive IMD treatment induces oxidative stress. Nonetheless, by increasing the appropriate levels of exogenous Mel, the photosynthesis of cucumber under IMD treatment reached the control levels, effectively removing reactive oxygen species. Furthermore, the content and ratio of ascorbate (AsA) and glutathione (GSH) were decreased under IMD treatment; Mel treatment enhanced the AsA/DHA and GSH/GSSG ratios, as well as the activities of MDHAR, DHAR and GR, suggesting that Mel could alleviate oxidative stress of cucumber treated with IMD by regulating the ascorbic acid-glutathione cycle. Importantly, IMD degradation rate and glutathione S-transferase (GST) activity increased after Mel treatment. The levels of transcripts encoding antioxidant enzymes GPX and GST (GST1,2 and 3) were also increased, indicating that Mel accelerated IMD degradation. These results suggest that Mel plays an important role in the detoxification of IMD by promoting GST activity and transcription and the AsA-GSH cycle, thus providing an approach for plants to reduce IMD residue through the plant's own detoxification mechanism.

Roles of CYP3A4, CYP3A5 and CYP2C8 drug-metabolizing enzymes in cellular cytostatic resistance.

Metabolic deactivation by cytochrome P450 (CYP) is considered a potential mechanism of anticancer drug resistance. However, this hypothesis is predominantly based on indirect pieces of evidence and/or is influenced by interfering factors such as the use of multienzymatic models. Thus, an experimental approach for its verification is needed. In the present work, we employed HepG2 cells transduced with CYP enzymes involved in docetaxel, paclitaxel and vincristine metabolism to provide mechanistic evidence on their possible roles in resistance to these chemotherapeutic agents. Using MTT proliferation tests, we showed that overexpression of CYP3A4 resulted in decreased antiproliferative activity of 1 µM docetaxel (by 11.2, 23.2 and 22.9% at 24, 48 and 72 h intervals, respectively), while the sensitivity of CYP3A4-transduced cells was restored by co-administration of ketoconazole. Paclitaxel exhibited differential efficacy in CYP2C8- and empty vector-transduced cells (significant differences between 10.9 and 24.4% for 0.01, 0.1 and 1 µM concentrations), but neither montelukast nor clotrimazole was capable of affecting this asymmetry. Finally, the pharmacological activity of vincristine was not influenced by CYP3A4 or CYP3A5 overexpression. In the follow-up caspase activation assays, docetaxel was confirmed to be a victim of CYP3A4-mediated resistance, which is, at least partly, brought by impaired activation of caspases 3/7, 8 and 9. In summary, our data demonstrate that CYP3A4-mediated metabolic deactivation of docetaxel might represent a significant mechanism of pharmacokinetic resistance to this drug. In contrast, the possible role of CYPs in resistance to paclitaxel and vincristine has been disconfirmed. Importantly, the expression of CYP3A4 in HepG2_CYP3A4 cells is comparable to that in primary hepatocytes and HepaRG cells, which suggests that our results might be relevant for in vivo conditions, e.g., for hepatocellular carcinoma. Thus, our data may serve as a valuable in vitro background for future in vivo studies exploring the area of intratumoural metabolism-based drug resistance.

Transcriptomic responses to aluminum stress in tea plant leaves.

Tea plant (Camellia sinensis) is a well-known Al-accumulating plant, showing a high level of aluminum (Al) tolerance. However, the molecular mechanisms of Al tolerance and accumulation are poorly understood. We carried out transcriptome analysis of tea plant leaves in response to three different Al levels (0, 1, 4 mM, for 7 days). In total, 794, 829 and 585 differentially expressed genes (DEGs) were obtained in 4 mM Al vs. 1 mM Al, 0 Al vs. 1 mM Al, and 4 mM Al vs. 0 Al comparisons, respectively. Analysis of genes related to polysaccharide and cell wall metabolism, detoxification of reactive oxygen species (ROS), cellular transport, and signal transduction were involved in the Al stress response. Furthermore, the transcription factors such as zinc finger, myeloblastosis (MYB), and WRKY played a critical role in transcriptional regulation of genes associated with Al resistance in tea plant. In addition, the genes involved in phenolics biosynthesis and decomposition were overwhelmingly upregulated in the leaves treated with either 0 Al and 4 mM Al stress, indicating they may play an important role in Al tolerance. These results will further help us to understand mechanisms of Al stress and tolerance in tea plants regulated at the transcriptional level.

Cancer-Specific Targeting of Taurine-Upregulated Gene 1 Enhances the Effects of Chemotherapy in Pancreatic Cancer.

Overcoming drug resistance is one of the biggest challenges in cancer chemotherapy. In this study, we examine whether targeting the long noncoding RNA taurine upregulated gene 1 (TUG1) could be an effective therapeutic approach to overcome drug resistance in pancreatic ductal adenocarcinoma (PDAC). TUG1 was expressed at significantly higher levels across 197 PDAC tissues compared with normal pancreatic tissues. Overall survival of patients with PDAC who had undergone 5-FU-based chemotherapy was shorter in high TUG1 group than in low TUG1 group. Mechanistically, TUG1 antagonized miR-376b-3p and upregulated dihydropyrimidine dehydrogenase (DPD). TUG1 depletion induced susceptibility to 5-FU in BxPC-3 and PK-9 pancreatic cell lines. Consistently, the cellular concentration of 5-FU was significantly higher under TUG1-depleted conditions. In PDAC xenograft models, intravenous treatment with a cancer-specific drug delivery system (TUG1-DDS) and 5-FU significantly suppressed PDAC tumor growth compared with 5-FU treatment alone. This novel approach using TUG1-DDS in combination with 5-FU may serve as an effective therapeutic option to attenuate DPD activity and meet appropriate 5-FU dosage requirements in targeted PDAC cells, which can reduce the systemic adverse effects of chemotherapy. SIGNIFICANCE: Targeting TUG1 coupled with a cancer-specific drug delivery system effectively modulates 5-FU catabolism in TUG1-overexpressing PDAC cells, thus contributing to a new combinatorial strategy for cancer treatment. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/7/1654/F1.large.jpg.

ABC transporter Pdr5 is required for cantharidin resistance in Saccharomyces cerevisiae.

Cantharidin is a potent anti-cancer drug and is known to exert its cytotoxic effects in several cancer cell lines. Although we have ample knowledge about its mode of action, we still know a little about cantharidin associated drug resistance mechanisms which dictates the efficacy and cytotoxic potential of this drug. In this direction, in the present study we employed Sacharomyces cerevisiae as a model organism and screened mutants of pleiotropic drug resistance network of genes for their susceptibility to cantharidin. We show that growth of pdr1Δ and pdr1Δpdr3Δ was severely reduced in presence of cantharidin whereas that of pdr3Δ remain unaffected when compared to wildtype. Loss of one of the PDR1 target genes PDR5, encoding an ABC membrane efflux pump, rendered the cells hypersensitive whereas overexpression of it conferred resistance. Additionally, cantharidin induced the upregulation of both PDR1 and PDR5 genes. Interestingly, pdr1Δpdr5Δ double deletion mutants were hypersensitive to cantharidin showing a synergistic effect in its cellular detoxification. Furthermore, transcriptional activation of PDR5 post cantharidin treatment was majorly dependent on the presence of Pdr1 and less significantly of Pdr3 transcription factors. Altogether our findings suggest that Pdr1 acts to increase cantharidin resistance by elevating the level of Pdr5 which serves as a major detoxification safeguard under CAN stress.

Dietary iso-α-acids prevent acetaldehyde-induced liver injury through Nrf2-mediated gene expression.

Acetaldehyde is the major toxic metabolite of alcohol (ethanol) and enhances fibrosis of the liver through hepatic stellate cells. Additionally, alcohol administration causes the accumulation of reactive oxygen species (ROS), which induce hepatocyte injury-mediated lipid peroxidation. Iso-α-acids, called isohumulones, are bitter acids in beer. The purpose of this study was to investigate the protective effects of iso-α-acids against alcoholic liver injury in hepatocytes in mice. C57BL/6N mice were fed diets containing isomerized hop extract, which mainly consists of iso-α-acids. After 7 days of feeding, acetaldehyde was administered by a single intraperitoneal injection. The acetaldehyde-induced increases in serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels were suppressed by iso-α-acids intake. Hepatic gene expression analyses showed the upregulation of detoxifying enzyme genes, glutathione-S-transferase (GST) and aldehyde dehydrogenase (ALDH). In vitro, iso-α-acids upregulated the enzymatic activities of GST and ALDH and induced the nuclear translocation of nuclear factor-erythroid-2-related factor 2 (Nfe2l2; Nrf2), a master regulator of antioxidant and detoxifying systems. These results suggest that iso-α-acid intake prevents acetaldehyde-induced liver injury by reducing oxidative stress via Nrf2-mediated gene expression.

Evaluation of hepatic drug-metabolism for glioblastoma using liver-brain chip.

Glioma is one of the most aggressive and highly fatal diseases with an extremely poor prognosis. Considering the poor clinical response to therapy in glioma, it is urgent to establish an in vitro model to facilitate the screening and assessment of anti-brain-tumor drugs. The blood-brain barrier (BBB), as well as liver metabolism plays an important role in determining the pharmacological activity of many anti-brain-tumor drugs. In this work, we designed a multi-interface liver-brain chip integrating co-culture system to assess hepatic metabolism dependent cytotoxicity of anti-brain-tumor drug in vitro. This microdevice composed of three microchannels which were separated by porous membrane and collagen. HepG2 and U87 cells were cultured in separated channels as mimics of liver and glioblastoma. Brain microvascular endothelial cells (BMECS) and cerebral astrocytes were co-cultured on collagen to mimic the brain microvascular endothelial barrier. Three common anti-tumor drugs, paclitaxel (PTX), capecitabine (CAP) and temozolomide (TMZ), were evaluated on this chip. In integrated liver-brain chip, liver enhanced the cytotoxicity of CAP on U87 cells by 30%, but having no significant effect on TMZ. The BBB decreased the cytotoxicity of PTX by 20%, while no significant effects were observed on TMZ and CAP, indicating the importance of liver metabolism and blood-brain barrier on the evaluation of anti-brain-tumor drugs. This work provides a biomimetic liver-brain model to mimic the physiological and pharmacological processes in vitro and presents a simple platform for long-term cell co-culture, drug delivery and metabolism, and real-time analysis of drug effects on brain cancer.

Gold Nanoparticles Perturb Drug-Metabolizing Enzymes and Antioxidants in the Livers of Male Rats: Potential Impact on Drug Interactions.

BACKGROUND AND AIM: With the wide applications of chitosan and gold nanoparticles in drug delivery and many consumer products, there is limited available information about their effects on drug-metabolizing enzymes (DMEs). Changes in DMEs could result in serious drug interactions. Therefore, this study aimed to investigate the effects of exposure to chitosan or gold nanoparticles on hepatic Phase I and II DMEs, liver function and integrity, oxidative damage and liver architecture in male rats. METHODS: Animals were divided into three equal groups: a control group, a group treated with chitosan nanoparticles (200 mg/kg, 50±5 nm) and a group treated with gold nanoparticles (4 mg/kg, 15±5 nm). Rats were orally administered their respective doses daily for 10 days. RESULTS: Both chitosan and gold nanoparticles decreased the body weights by more than 10%. Gold nanoparticles reduced the activities of antioxidants (superoxide dismutase and catalase), and reduced glutathione level and elevated the malondialdehyde level in the liver. Gold nanoparticles caused significant reductions in CYP1A1, CYP2E1, quinone oxidoreductase1, and glutathione S-transferase and elevated CYP2D6 and N-acetyl transferase2. Chitosan elevated CYP2E1 and CYP2D6 and reduced UDP-glucuronosyltransferase 1A1. Both nanoparticles disturbed the architecture of the liver, but the deleterious effects after gold nanoparticles treatment were more prominent. CONCLUSION: Taken together, gold nanoparticles severely perturbed the DMEs and would result in serious interactions with many drugs, herbs, and foods.

Combined effects of environmentally relevant concentrations of diclofenac and cadmium on Chironomus riparius larvae.

The nonsteroidal anti-inflammatory drug diclofenac (DCF) is considered a contaminant of emerging concern. DCF can co-exist with heavy metals in aquatic environments, causing unexpected risks to aquatic organisms. This study aimed to assess the combined effects of DCF and cadmium (Cd) at environmentally relevant concentrations on the bioconcentration and status of oxidative stress and detoxification in Chironomus riparius larvae. The larvae were exposed to DCF (2 and 20 µg L-1) and Cd (5 and 50 µg L-1) alone or in mixtures for 48 h. The combined exposure to DCF and Cd was found to reciprocally facilitate the accumulation of each compound in larvae compared with single exposures. As indicated by the antioxidant enzyme activities, reduced glutathione levels, and malondialdehyde contents, the low concentration of the mixture (2 µg L-1 DCF + 5 µg L-1 Cd) did not alter the oxidative stress status in larvae, while the high concentration of the mixture (20 µg L-1 DCF + 50 µg L-1 Cd) induced stronger oxidative damage to larvae compared with single exposures. The expression levels of eight genes (CuZnSOD, MnSOD, CAT, GSTd3, GSTe1, GSTs4, CYP4G, and CYP9AT2) significantly decreased due to the high concentration of the mixture compared with single exposures in most cases. Overall, the results suggest that the mixture of DCF and Cd might exert greater ecological risks to aquatic insects compared with their individual compounds.

Studies of selenium and arsenic mutual protection in human HepG2 cells.

Hundreds of millions of people worldwide are exposed to unacceptable levels of carcinogenic inorganic arsenic. Animal models have shown that selenium and arsenic are mutually protective through the formation and elimination of the seleno-bis(S-glutathionyl) arsinium ion [(GS)2AsSe]-. Consistent with this, human selenium deficiency in arsenic-endemic regions is associated with arsenic-induced disease, leading to the initiation of human selenium supplementation trials. In contrast to the protective effect observed in vivo, in vitro studies have suggested that selenite increases arsenite cellular retention and toxicity. This difference might be explained by the rapid conversion of selenite to selenide in vivo. In the current study, selenite did not protect the human hepatoma (HepG2) cell line against the toxicity of arsenite at equimolar concentrations, however selenide increased the IC50 by 2.3-fold. Cytotoxicity assays of arsenite + selenite and arsenite + selenide at different molar ratios revealed higher overall mutual antagonism of arsenite + selenide toxicity than arsenite + selenite. Despite this protective effect, in comparison to 75Se-selenite, HepG2 cells in suspension were at least 3-fold more efficient at accumulating selenium from reduced 75Se-selenide, and its accumulation was further increased by arsenite. X-ray fluorescence imaging of HepG2 cells also showed that arsenic accumulation, in the presence of selenide, was higher than in the presence of selenite. These results are consistent with a greater intracellular availability of selenide relative to selenite for protection against arsenite, and the formation and retention of a less toxic product, possibly [(GS)2AsSe]-.

Bioactive compounds from Aspergillus niger extract enhance the antioxidant activity and prevent the genotoxicity in aflatoxin B1-treated rats.

This study aimed to identify the bioactive compounds of the ethyl acetate extract of Aspergillus niger SH2-EGY using GC-MS and to evaluate their protective role against aflatoxin B1 (AFB1)-induced oxidative stress, genotoxicity and cytotoxicity in rats. Six groups of male Sprague-Dawley rats were treated orally for 4 weeks included the control group, AFB1-treated group (80 µg/kg b.w); fungal extract (FE)-treated groups at low (140) or high dose (280) mg/kg b.w and the groups treated with AFB1 plus FE at the two tested doses. The GC-MS analysis identified 26 compounds. The major compounds found were 1,2,3,4,6-Penta-trimethylsilyl Glucopyranose, Fmoc-L-3-(2-Naphthyl)-alanine, D-(-)-Fructopyranose, pentakis (trimethylsilyl) ether, bis (2-ethylhexyl) phthalate, trimethylsilyl ether-glucitol, and octadecanamide, N-(2- methylpropyl)-N-nitroso. The in vivo results showed that AFB1 significantly increased serum ALT, AST, creatinine, uric acid, urea, cholesterol, triglycerides, LDL, carcinoembryonic antigen, alpha-fetoprotein, interleukin-6, Malondialdehyde, nitric oxide, Bax, caspase-3 and P53 mRNA expression, chromosomal aberrations and DNA fragmentation. It decreased serum TP, albumin, HDL, Bcl-2 mRNA expression, hepatic and renal TAC, SOD and GPx content and induced histological changes in the liver and kidney. FE prevented these disturbances in a dosage-dependent manner. It could be concluded that A. niger SH2-EGY extract is safe a promising agent for pharmaceutical and food industries.

Mitochondrial Respiratory Inhibition Promoted by Pyraclostrobin in Fungi is Also Observed in Honey Bees.

There is no use restriction associated with bees for many fungicides used in agriculture; however, this does not always mean that these pesticides are harmless for these nontarget organisms. We investigated whether the fungicide pyraclostrobin, which acts on fungal mitochondria, also negatively affects honey bee mitochondrial bioenergetics. Honey bees were collected from 5 hives and anesthetized at 4 °C. The thoraces were separated, and mitochondria were isolated by grinding, filtering, and differential centrifugation. An aliquot of 0.5 mg of mitochondrial proteins was added to 0.5 mL of a standard reaction medium with 4 mM succinate (complex II substrate) plus 50 nM rotenone (complex I inhibitor), and mitochondrial respiration was measured at 30 °C using a Clark-type oxygen electrode. Mitochondrial membrane potential was determined spectrofluorimetrically using safranin O as a probe, and adenosine triphosphate (ATP) synthesis was determined by chemiluminescence. Pyraclostrobin at 0 to 50 µM was tested on the mitochondrial preparations, with 3 repetitions. Pyraclostrobin inhibited mitochondrial respiration in a dose-dependent manner at concentrations of 10 µM and above, demonstrating typical inhibition of oxidative phosphorylation. Pyraclostrobin also promoted a decline in the mitochondrial membrane potential at doses of 5 µM and above and in ATP synthesis at 15 µM and above. We conclude that pyraclostrobin interferes with honey bee mitochondrial function, which is especially critical for the energy-demanding flight activity of foraging bees. Environ Toxicol Chem 2020;39:1267-1272. © 2020 SETAC.

Glutathione-Allylsulfur Conjugates as Mesenchymal Stem Cells Stimulating Agents for Potential Applications in Tissue Repair.

The endogenous gasotransmitter H2S plays an important role in the central nervous, respiratory and cardiovascular systems. Accordingly, slow-releasing H2S donors are powerful tools for basic studies and innovative pharmaco-therapeutic agents for cardiovascular and neurodegenerative diseases. Nonetheless, the effects of H2S-releasing agents on the growth of stem cells have not been fully investigated. H2S preconditioning can enhance mesenchymal stem cell survival after post-ischaemic myocardial implantation; therefore, stem cell therapy combined with H2S may be relevant in cell-based therapy for regenerative medicine. Here, we studied the effects of slow-releasing H2S agents on the cell growth and differentiation of cardiac Lin- Sca1+ human mesenchymal stem cells (cMSC) and on normal human dermal fibroblasts (NHDF). In particular, we investigated the effects of water-soluble GSH-garlic conjugates (GSGa) on cMSC compared to other H2S-releasing agents, such as Na2S and GYY4137. GSGa treatment of cMSC and NHDF increased their cell proliferation and migration in a concentration dependent manner with respect to the control. GSGa treatment promoted an upregulation of the expression of proteins involved in oxidative stress protection, cell-cell adhesion and commitment to differentiation. These results highlight the effects of H2S-natural donors as biochemical factors that promote MSC homing, increasing their safety profile and efficacy after transplantation, and the value of these donors in developing functional 3D-stem cell delivery systems for cardiac muscle tissue repair and regeneration.

The comparison of transcriptomic response of green microalga Chlorella sorokiniana exposure to environmentally relevant concentration of cadmium(II) and 4-n-nonylphenol.

The transcriptomic response of green microalga Chlorella sorokiniana exposure to environmentally relevant concentration of cadmium(II) (Cd) and 4-n-nonylphenol (4-n-NP) was compared in the present study. Cd and 4-n-NP exposure showed a similar pattern of dys-regulated pathways. The photosystem was affected due to suppression of chlorophyll biosynthesis via down-regulation of Mg-protoporphyrin IX chelatase subunit ChlD (CHLD) and divinyl chlorophyllide a 8-vinyl-reductase (DVR) in Cd group and via down-regulation of DVR in 4-n-NP group. Furthermore, the reactive oxygen species (ROS) could be induced through down-regulation of solanesyl diphosphate synthase 1 (SPS1) and homogentisate phytyltransferase (HPT) in Cd group and via down-regulation of HPT in 4-n-NP group. Additionally, Cd and 4-n-NP would both cause the dys-regulation of carbohydrate metabolism and protein synthesis. On the other hand, there are some different responses or detoxification mechanism of C. sorokiniana to 4-n-NP stress compared to Cd exposure. The increased ROS would cause the DNA damage and protein destruction in Cd exposure group. Simultaneously, the RNA transcription was dys-regulated and a series of changes in gene expressions were observed. This included lipid metabolism, protein modification, and DNA repair, which involved in response of C. sorokiniana to Cd stress or detoxification of Cd. For 4-n-NP exposure, no effect on lipid metabolism and DNA repair was observed. The nucleotide metabolism including pyrimidine metabolism and purine metabolism was significantly up-regulated in the 4-n-NP exposure group, but not in the Cd exposure group. In addition, 4-n-NP would induce the ubiquitin-mediated proteolysis and proteasomal degradation to diminish the misfolded protein caused by ROS and down-regulation of heat shocking protein 40. In sum, the Cd and 4-n-NP could cause the same toxicological effects via the common pathways and possess similar detoxification mechanism. They also showed different responses in nucleotide metabolism, lipid metabolism, and DNA repair.

Role of Drug-metabolizing Enzymes in Cancer and Cancer Therapy.

BACKGROUND: Cancer is one of the most serious diseases threatening human health with high morbidity and mortality in the world. For the treatment of cancer, chemotherapy is one of the most widely used strategies, for almost all kinds of tumors and diverse stages of tumor development. The efficacy of chemotherapy not only depends on the activity of the drug administrated but also on whether the compound could reach the effective therapeutic concentration in tumor cells. Therefore, expression and activity of drug-metabolizing enzymes (DMEs) in tumor tissues and metabolic organs of cancer patients are important for the dispositional behavior of anticancer drugs as well as the clinical response of chemotherapy. METHODS: This review summarizes the recent advancement of the DMEs expression and activity in various cancers, as well as the potential regulatory mechanisms of major DMEs in cancer and cancer therapy. RESULTS: Compared to normal tissues, expression and activity of major DMEs are significantly dysregulated in patients by various factors including epigenetic modification, ligand-activated transcriptional regulation and signaling pathways. Additionally, DMEs play an important role in anticancer drug efficacy, chemoresistance as well as the activation of prodrugs. CONCLUSION: This review reinforces a more comprehensive understanding of DMEs in cancer and cancer therapy, and provides more opportunities for cancer therapy.