Inhibition Of Glutathione S-transferase Activity Research Articles

Polycyclic aromatic hydrocarbon (PAH) activation of AhR inhibits glutathione S-transferase (GST) activity in primary human small airway epithelial cells (hSAECs)

Background: PAH represents the organic phase of particulate matter (PM) in air pollution emitted via the incomplete combustion of various compounds (i.e. diesel fuel, wood, and plastics). It is widely regarded that PAH activation of aryl hydrocarbon receptor (AhR) plays an important role in pro-inflammatory signaling, however the mechanisms remain unknown. In contrast- endogenous AhR agonists, such as 6-Formylindolo[3,2-b]carbazole (FICZ), exert pleiotropic protective effects against inflammation, fibrosis, and oxidative stress. The objective of our research program is to delineate the differences between activation of AhR by PAH (with adverse effects) compared to its beneficial homeostatic activation by endogenous FICZ. PM may induce oxidative stress in part through inhibition of antioxidant defense mechanisms. GSTs are a superfamily of enzymes that plays an important role in protecting cells from oxidative damage (by conjugating reduced glutathione to substrates under pro-oxidizing conditions). We hypothesize that PAH activation of AhR contributes to oxidative stress via inhibition of GST activity in primary hSAECs. Methods: Primary hSAECs were subjected to varying dosages of PAH [0, 250, 500, 1000, 5000 ng/mL] to model airborne PM exposure levels, and to increasing doses of FICZ [0, 0.50, 10, 100, 1000 nM] to model normal and excessively high endogenous ligand activation of AhR. We determined the relative potencies of PAH and FICZ by measuring CYP1A1 and CYP1B1 gene expression. Oxidative stress was measured using DHE-labeling and GST activity was measured with enzymatic kits (Sigma Aldrich). AhR activity was pharmacologically inhibited using 10μM CH223191. Statistical analysis was conducted using SigmaPlot 14.0 (Systat Software). Results: PAH and FICZ maximally stimulate hSAEC CYP1A1 and CYP1B1 expression at 1000ng/mL and 100nM, respectively, and are therefore defined as equipotent concentrations used in our studies. In a representative experiment in hSAECs, PAH (1000ng/mL) significantly increased hSAEC oxidative stress vs FICZ (100nM) n=6, p<0.01), while significantly decreasing GST specific activity vs equipotent FICZ activation of AhR in primary hSAECs from a 66-year-old female without lung disease (26.6±0.874 vs 28.3±.611 nmol/mL/min); n=6 from 2 independent assays; p=0.032. Similar differential effects of PAH vs FICZ activation of AhR on GST were seen in additional experiments with male hSAECs and Calu-3 cells (male). Importantly, CH223191 inhibition of GST enzymatic activity suggests that AhR plays an important role in protecting cells from oxidative stress. Conclusions: Our studies indicate that airborne PAH exposure can have detrimental health effects on the lung via aberrant AhR signaling. Significance: Understanding activity of AhR signaling can leads to novel therapeutic options for redox disorders in the lung. Funding: VA 5I01BX001777-05 (RP) and R01HL137033 (MH). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The Effect and Mechanism of Curcumin Combined with Carboplatin Chemotherapy Promoting on Apoptosis of Lung Cancer HCC827 Cells.

Objective To investigate the effect and mechanism of curcumin (CUR) killing lung cancer HCC827 cell spheres. Method HCC827 cell spheres were cultured in serum-free medium, and the protein expression of CD133, SOX2, EpCAM, and ABCG2 was detected by western blot. MTT was used to evaluate the cell viability of HCC827 cell spheres and HCC827 cell after they were treated by 1, 2, 5, 10, and 20 mg/mL carboplatin (CBP) for 48 h. The inhibitory effects of 10 μM, 50 μM, 100 μM, and 200 μM CUR on GST (glutathione S-transferase) activity in HCC827 cell spheres were determined by colorimetry. The flow cytometry (FCM), western blot, qPCR, luciferase assay, and microscopy were used to detect the ROS levels, cell pelletization ability, β-catenin, SOX2, and ABCG2 mRNA and the promoter activity of β-catenin upon of HCC827 cell spheres treated with 200 μM CUR for 48 h. The HCC827 cell spheres were infected with β-catenin adenovirus, and then cells were treated with 200 μM CUR (and/or no 5 mg/mL CBP) for 24 h. The mRNA and protein expression of β-catenin, SOX2, and ABCG2 was detected by qPCR and western blot, and cell growth inhibition of HCC827 cell spheres was evaluated by MTT. Result The expression of stem cells marker CD133, SOX2, EpCAM, and drug resistance-related gene ABCG2 mRNA is higher in HCC827 cell spheres, and HCC827 cell spheres resisted the killing effect of difference doses of CBP. The activity of GST of HCC827 cell spheres was inhibited by 10 μM, 50 μM, 100 μM, and 200 μM CUR. It was a dose-dependent manner. After 200 μM CUR had treated HCC827 cell spheres for 48 h, the level of ROS was significantly increased (P < 0.05), and the mRNA and protein expression of β-catenin, SOX2, and ABCG2 and promoter activity of β-catenin were notably decreased (P < 0.05), compared to the control group. Furthermore, the formed-sphere ability of HCC827 sphere was inhibited after cells were treated with 200 μM CUR. 200 μM CUR could suppress the proliferation of HCC827 cell spheres and induced cell apoptosis. The proliferation of HCC827 cell spheres was significantly inhibited, and cell apoptosis rate was increased by 200 μM CUR combined with 5 mg/mL CBP than by 200 μM CUR alone. Upregulation of β-catenin by adenovirus partly reversed the effect of CUR inhibition of the expression of β-catenin, SOX2, and ABCG2, compared to empty vector adenovirus group. Additionally, overexpression of β-catenin significantly remitted the inhibitory effect of 200 μM CUR combined with 5 mg/mL CBP on the proliferation of HCC827 cell spheres. Conclusion CUR inhibited the cell proliferation and stem cell trait and induced apoptosis in HCC827 cell spheres by the inhibition of GST activity and β-catenin expression. CUR is expected to become a treatment for lung cancer and lung cancer stem cells.

Chemical composition of essential oils from Thymus mongolicus, Cinnamomum verum, and Origanum vulgare and their acaricidal effects on Haemaphysalis longicornis (Acari: Ixodidae)

Chemical acaricides are mainly used in traditional tick control, which leads to the emergence of tick resistance and concurrently results in environmental pollution. In the present study, the chemical constituents of essential oils (EOs) from Thymus mongolicus, Cinnamomum verum, and Origanum vulgare was analyzed, and their potential application was evaluated to control the vector tick Haemaphysalis longicornis, which is widely distributed over vast areas of Eurasia, Australia, and New Zealand. Gas chromatography-mass spectrometry analysis revealed that the phenols thymol and carvacrol accounted for 34.66% and 75.72% of the EOs of T. mongolicus and O. vulgare, respectively, whereas trans-cinnamaldehyde (49.42%) was the main constituent of C. verum EO. Immersion tests showed that the EOs of C. verum and O. vulgare had significant acaricidal activity against larval H. longicornis, with the 50% lethal concentration (LC50) being 16.07 and 18.02 mg/mL, respectively, and the 95% lethal concentration (LC95) being 120.37 and 130.09 mg/mL, respectively. The EOs of O. vulgare and T. mongolicus showed significant acaricidal activity against unfed adult H. longicornis, with LC50 being 43.50 and 44.21 mg/mL, respectively, and LC95 being 113.66 and 137.99 mg/mL, respectively. The fumigant toxicity test showed significant acaricidal activity of the three EOs against both unfed and engorged nymphal and adult H. longicornis. Enzyme assays revealed that the EOs of both C. verum and O. vulgare significantly inhibited glutathione S-transferase activity (P < 0.05). In contrast, the activities of carboxylesterase and multifunction oxidases were significantly inhibited by EOs extracted from all three plants (P < 0.05). Taken together, these findings suggest that plant EOs may serve as an environment-friendly alternative for synthetic acaricides in future tick control.

Tissue distribution and sublethal effects of imidacloprid in the South American grayish baywing (Agelaioides badius)

The neonicotinoids are globally used insecticides, which have been shown to cause negative impacts on birds. The current study aimed to evaluate the distribution of the neonicotinoid imidacloprid (IMI) in the tissues of a songbird and identify related physiological effects. Adults of the grayish baywing (Agelaioides baduis) were administered with a single dose of 35 mg IMI/kg, and the IMI concentration was evaluated in liver, kidney and plasma at 4, 12, 24, and 48 h after dosing. At the same time points, effects on hematological, genetic and enzymatic parameters were assessed. Results showed that IMI was absorbed before 4 h, and eliminated at 48 h, in every tissue, and the highest concentrations were detected in plasma. Baywings showed intoxication signs and reduced mobility within the first 5 min post-dosing. Hematological parameters: red blood cells, packed cell volume, hemoglobin, and their derived indices exhibited a transient elevation 24 h after dosing, which coincided with maximum concentrations of IMI in the tissues. No effects were observed on the genotoxicity parameters evaluated: micronuclei and comet assay. Treated birds exhibited an alteration of cholinesterases activity in the muscle and plasma, and of glutathione-S-transferase (GST) activity in the plasma, brain, liver, and muscle. Based on the results obtained, the combined detection of IMI and inhibition of GST activity in the plasma is suggested as a non-lethal biomarker of IMI exposure in wild birds. As efficient field monitoring depends on the availability of proven biomarkers, the current study provides valuable tools for bird conservation in agroecosystems.

Molecular and biochemical evaluation of effects of malathion, phenanthrene and cadmium on Chironomus sancticaroli (Diptera: Chironomidae) larvae

In-vitro effects of sub-lethal concentrations of malathion, phenanthrene (Phe) and cadmium (Cd) were tested on Chironomus sancticaroli larvae in acute bioassays by measuring biochemical and molecular parameters. Malathion was evaluated at 0.001, 0.0564 and 0.1006 mg L-1; Phe at 0.0025, 1.25 and 2.44 mg L-1; and Cd at 0.001, 3.2 and 7.4 mg L-1. The recovery test carried out at the highest concentration of each compound showed that survival of larvae exposed to Phe ranged from 4% to 5%, while the effects of malathion and Cd were irreversible, not allowing the emergence of adults. Results showed that malathion and Cd inhibited AChE, EST-α and ES-β activities at the two highest concentrations. Phe at 0.0025, 1.25 and 2.44 mg L-1; and Cd at 3.2 and 7.4 mg L-1 inhibited glutathione S-transferase activity. Oxidative stress was exclusively induced by the lowest concentration of malathion considering SOD activity once CAT was unaffected by the stressors. Lipid peroxidation was registered exclusively by malathion at the two highest concentrations, and total hemoglobin content was only reduced by Cd at the two highest concentrations. The relationship among biochemical results, examined using the PCA, evidenced that malathion and Cd concentrations were clustered into two groups, while Phe only formed one group. Four hemoglobin genes of C. sancticaroli were tested for the first time in this species, with Hemoglobin-C being upregulated by malathion. The toxicity ranking was malathion > Phe > Cd, while biochemical and molecular results showed the order malathion > Cd > Phe. Our results highlight the importance of combining different markers to understand the effects of the diverse compounds in aquatic organisms.

Oxidative stress and genotoxicity in Rhinella arenarum (Anura: Bufonidae) tadpoles after acute exposure to Ni-Al nanoceramics

The employ of nanomaterials (NMs) has exponentially grown due to the large number of technological advances in industrial, pharmaceutical and medical areas. That is the case of alumina (Al) nanoparticles which are extensively employed as support in heterogeneous catalysis processes. However, these NMs can cause great toxicity because of their ubiquitous properties, such as extremely small size and high specific surface area. So, it is required to assess the potential deleterious effects of these NMs on living organisms. In the present study, we analyze the oxidative stress and genotoxic potential of a nanoceramic catalyst Ni/<gamma>-Al2O3 (NC) and the NMs involved in their synthesis, <gamma>-Al2O3 support (SPC) and NiO/<gamma>-Al2O3 precursor (PC) on Rhinella arenarum larvae. Biomarkers of oxidative stress and genotoxic damage were measured in tadpoles exposed to 5 and 25 mg/L of each NMs for 96 h. The results indicated an inhibition of catalase activity in tadpoles exposed to both concentrations of PC and to 25 mg/L of SPC and NC. Moreover, both exposure concentrations of PC and NC significantly inhibited superoxide dismutase activity. Exposure to the three NMs caused inhibition of glutathione S-transferase activity, but there were no significant variations in reduced glutathione levels. Oxidative stress damage (lipid peroxidation) was observed in tadpoles treated with 25 mg/L PC, while the other treatments did not produce alterations. The MNs frequency significantly increased in larvae exposed to 25 mg/L PC indicating irreversible genotoxic damage. The results show that these NMs exert genotoxic effects and antioxidant defense system disruption in R. arenarum larvae.

Activity of Noni Fruit (Morinda citrifolia L.) Ethanolic Extract on Class mu Glutation S-Transferase of Lung Rat

One of the main modalities of cancer treatment is chemotherapy, which uses chemicals that are generally electrophilic. These xenobiotic compounds sometimes does not produce effective response due to activity of glutathione S-transferase (GST) which inactivate the xenobiotics. Several natural phenolic compounds were reported to inhibit GST activity in vitro. Noni fruit (Morinda citrifolia L.) which contains flavonoids and other phenolic compounds such as scopoletin and morindon is proposed to interfere GST activity. This study aimed to analyze the effect of ethanolic extract of Noni fruit in vivo on GST activity in lung rat using 1,2-dichloro-4-nitrobenzene (DCNB). This substrate is a specific for class mu GST. First, rats were administered with ethanolic extract of Noni and dimethylbenz(α)anthracene (DMBA) for two weeks. The cytosolic fraction of lung was isolated then the GST activity was determined by simple kinetic program which was automatically calculated using spectrophotometer. The results showed that ethanolic extract of Noni in 1 and 5% (w/v) of concentration induced class mu GST activity, whereas 10% (w/v) of concentration inhibited class mu GST activity. After a treatment with DMBA, all tested concentrations of ethanolic extract of Noni inhibited class mu GST activity of lung rat significantly. These results indicated that Noni fruit extract can be further developed as a supportive agent of a chemotherapy drug.Keywords: DMBA, GST, Morinda citrifolia L., spectrophotometer.

Effects of chronic exposures of selected heavy metals on the glutathione S-transferase activity of freshwater snails Lymnaea natalensis in Zimbabwe

The effect of the heavy metals (cadmium, copper, mercury and lead) on snail glutathione S-transferase (GST) was investigated in 2015. Groups of Lymnaea natalensis snails were exposed to heavy metals for 28 days at concentrations reportedly found in the Mguza Dam. Water and food were changed daily. Samples were collected at days 1, 7, 14, 21 and 28 post exposure. Inhibition of GST activity, following cadmium exposures, ranged between 58 and 60%, with a decrease of 30% on day 28. When snails were exposed to copper, inhibition significantly decreased by 16%, 29%, 49% and 72% inhibition when tested on days 1, 7, 14 and 21, respectively. Inhibition on day 28 was 44%. Mercury exposures resulted in significant increases in GST inhibition, namely, 47%, 62% and 79% inhibition on days 1, 7 and 14, respectively. Inhibition on day 21 was 82%, whereas on day 28 it was significantly lower, at 29%. Concerning lead exposures, inhibition levels on day 1, 7 and 21 had mean inhibition of 60%. Inhibition on days 14 and 28 was significantly lower, with a mean inhibition of 30%. These results suggest that chronic exposures could inhibit GST activity for a certain period, after which inhibition is reduced, possibly as a result of adaptation.

MODIFICATION OF THE NANO TITANIUM BAND GAP REACTIONS OF SOLID DIOXIDE WITH (ANTIMONY) STIBIUM S AND ITS DETERMINATION PHOTOCATALYTIC ACTIVITY

This study includes preparation of nanoparticles of titanium dioxide using the sol-gel method and then, the band gap was modified by solid-state reaction with (Antimony) Stibium Sb reductive compounds. Modification reduced the band gap separating energy levels between valance band (VB) and conduction band (CB), therefore; facilitating the transfer of excited electrons from (VB) to (CB). Absorption of the energy from incident photons having the same or larger energy than the energy of the band gap promote the formation of the couple (electron-hole). The resulting (e--h+) couple acts to produce (OH.) radicals. OH. radicals having a high capability to inhibition of glutathione S-transferase activity in Aedes aegypti, mosquitoes that adsorbed on the surface of the photocatalytic TiO2 as a new model of eco-friendly insecticide.

Anatolıan sage Salvıa frutıcosa ınhıbıts cytosolıc glutathıone-s-transferase actıvıty and colon cancer cell prolıferatıon

Colorectal cancer is one of the leading causes of death worldwide, and healthy diet is critical in preventing it. This study intends to investigate phenolic profile, antioxidant, and anti-proliferative activity as well as Glutathione S-transferase (GST) inhibition activity of water extract of Salvia fruticosa Mill. (SF.) HPLC analysis revealed that the main bioactive ingredients of SF were rosmarinic and caffeic acid. Total phenolic and flavonoid contents were 425.1 mg GAE/g extract and 94.7 mg CE/g extract, respectively. SF exhibited high antioxidant activity through DPPH and ABTS assays. The extract exhibited time and dose-dependent anti-proliferative activity on colorectal adenocarcinoma cell lines (Caco-2 and HT-29). The extract also inhibited GST activity in a mixed-type manner, with an IC50 value of 6.8 µg/mL. To conclude, the results suggest that SF may be considered as a medicinal plant which can be used in theprevention and treatment of colon cancer, besides chemotherapy drugs.

Bioconcentration, behavioral, and biochemical effects of the non-steroidal anti-inflammatory drug diclofenac in Daphnia magna.

The non-steroidal anti-inflammatory drug (NSAID) diclofenac is one of the most frequently studied as well as controversially discussed pharmaceutically active drug on the subject of its relevance to the environment. This study was conducted to assess the bioconcentration potential of diclofenac and its behavioral and biochemical effects in Daphnia magna. The bioconcentration factors of diclofenac determined after 48h of aqueous exposure in Daphnia magna were 70.94 and 8.02 for the nominal exposure concentrations of 5 and 100μg/L, respectively. Diclofenac exposure obviously decreased the filtration and ingestion rates of the daphnids. A significant increase of the acetylcholinesterase activity that was observed in this study indicates that diclofenac might not have neurobehavioral toxicity in Daphnia magna. Significant induction of malondialdehyde content is an indication of overproduction of reactive oxygen species leading to oxidative damage in daphnids after diclofenac exposure. Moreover, significant inhibition of the superoxide dismutase, catalase, and glutathione reductase activities implies that the antioxidant defense system of Daphnia magna was overwhelmed. Also, significant inhibition of glutathione s-transferase activity might point to the fact that the enzyme was not capable to detoxify diclofenac in Daphnia magna. These findings indicate that diclofenac can accumulate and consequently stimulate behavioral and biochemical disturbances in Daphnia magna.

Inhibitory effects of ficus deltoidea extracts on UDP-glucuronosyltransferase and glutathione s-transferase drug-metabolizing enzymes

Background: Mas cotek or Ficus deltoidea is conventionally used to treat various diseases and often consumed with other medication and this may give rise to herb–drug interaction. The potential of F. deltoidea for interactions with drug-metabolizing enzymes of UDP-gucuronosytranserase (UGT) and glutathione S-transferase (GST) have not been investigated. Objective: We had evaluated the potential of methanol, ethanol, and aqueous extracts of F. deltoidea to cause UGT- and GST-mediated herb–drug interaction in vitro. Materials and Methods: The total phenolic content and total phenolic content were determined using modified colorimetric method. In the UGT study, para -nitrophenol (p -NP) was employed as a substrate to determine the UGT enzymes activity in rat liver microsomes (RLM) and human liver microsomes (HLM). For the GST study, 1-chloro-2,4-dinitrobenzene was employed as a substrate to determine GST activity in rat liver cytosolic fraction. Results: The total phenolic content in F. deltoidea extracts can be ranked as follows: Methanol extract > aqueous extract > ethanol extract, whereas the content of flavonoid compounds in F. deltoidea extracts can be ranked as: Methanol extract > ethanol extract > aqueous extract. Assessment using the UGT enzymes of RLM (IC50 [Half-maximal inhibitory concentration] = 881.40 ± 1.14 μg/mL) and HLM (IC50 =63.44 ± 1.20 μg/mL) showed that the methanol extract of F. deltoidea significantly inhibited p -NP glucuronidation compared with ethanol and aqueous extracts. For GST inhibition study, methanol extract strongly inhibited GST activity (IC50 =70.73 ± 1.07 μg/mL), whereas no IC50values were determined for ethanol and aqueous extracts. Conclusion: The methanol extract of F. deltoidea containing the highest flavonoid content highlights the possibility of herb–drug interaction through the modulation of p -NP UGT and GST activity.

Influence of sex on biomarkers of oxidative stress in the kidney, lungs, and liver of rabbits after exposure to diazinon

The present study evaluated the effect of two oral doses of the pesticide diazinon (25 and 125mg/kg bw) on four biochemical parameters (malondialdehyde and glutathione levels, and catalase and glutathione S-transferase enzymatic activities) in the kidney, lungs, and liver of rabbit after 10days of exposure. Malondialdehyde levels were significantly greater in exposed animals compared to controls, especially in the animals exposed to the higher dose of the pesticide. This result was reflected in the glutathione levels, which were significantly lower at that same higher dose. Catalase activity was also inhibited by the higher dose of the pesticide in all three organs analysed, whereas inhibition of glutathione S-transferase activity was only significant for the kidney and lungs. There were some slight differences between the sexes: taking the results for all three organs, the higher dose of diazinon resulted in a clearly significant inhibitory effect on the catalase activity and glutathione levels in males, and a significant enhancing effect on the malondialdehyde levels in females. These results help to confirm the interest of considering such endogenous factors in future ecotoxicological studies, and that the four biomarkers considered are suitable for reflecting the toxic effects of diazinon on rabbits, especially the effects related to oxidative stress. Graphical abstract.

Survival, Reproduction, Avoidance Behavior and Oxidative Stress Biomarkers in the Earthworm Octolasion cyaneum Exposed to Glyphosate.

The massive use of glyphosate (GLY) in several countries has increased the interest in investigating its potential adverse effects in non-target organisms. The aim of the present study was to assess the potential effects in survival and reproduction; avoidance behavior and oxidative stress under short-term (48h) and subchronic exposures (28days) to GLY in the earthworm Octolasion cyaneum. After 48h no significant changes in the behavior was observed. In addition, a lower catalase activity at 498μgGLYkg(-1) dry soil section relative to earthworms from the control section was obtained. After 28days of exposure inhibition of glutathione S-transferase activity was observed at 535μgGLYkg(-1) dry soil while no changes in the other endpoints were detected. These results indicate that environmentally relevant concentrations of GLY (up to 996µgGLYkg(-1) dry soil) did not exert a toxic effect to O. cyaneum.

Bioactivation of luteolin by tyrosinase selectively inhibits glutathione S-transferase

Glutathione S-transferase (GST) plays a significant role in the metabolism and detoxification of drugs used in treatment of melanoma, resulting in a decrease in drug efficacy. Tyrosinase is an abundant enzyme found in melanoma. In this study, we used a tyrosinase targeted approach to selectively inhibit GST. In the presence of tyrosinase, luteolin (10 μM) showed 87% GST inhibition; whereas in the absence of tyrosinase, luteolin led to negligible GST inhibition. With respect to GSH, both luteolin-SG conjugate and luteolin-quinone inhibited ≥90% of GST activity via competitive reversible and irreversible mixed mechanisms with Ki of 0.74 μM and 0.02 μM, respectively. With respect to CDNB, the luteolin-SG conjugate inhibited GST activity via competitive reversible mechanism and competitively with Ki of 0.58 μM, whereas luteolin-quinone showed irreversible mixed inhibition of GST activity with Ki of 0.039 μM. Luteolin (100 μM) inhibited GST in mixed manner with Ki of 53 μM with respect to GSH and non-competitively with respect to CDNB with Ki of 38 μM. Luteolin, at a concentration range of 5–80 μM, exhibited 78–99% GST inhibition in human SK-MEL-28 cell homogenate. Among the 3 species of intact luteolin, luteolin-SG conjugate, and luteoline-quinone, only the latter two have potential as drugs with Ki < 1 μM, which is potentially achievable in-vivo as therapeutic agents. The order of GST inhibition was luteolin-quinone >> luteolin-SG conjugate >>> luteolin. In summary, our results suggest that luteolin was bioactivated by tyrosinase to form a luteolin-quinone and luteolin-glutathione conjugate, which inhibited GST. For the first time, in addition to intracellular GSH depletion, we demonstrate that luteolin acts as a selective inhibitor of GST in the presence of tyrosinase. Such strategy could potentially be used to selectively inhibit GST, a drug detoxifying enzyme, in melanoma cells.

Fallopia japonica, a Natural Modulator, Can Overcome Multidrug Resistance in Cancer Cells.

Resistance of cancer cells to chemotherapy is controlled by the decrease of intracellular drug accumulation, increase of detoxification, and diminished propensity of cancer cells to undergo apoptosis. ATP-binding cassette (ABC) membrane transporters with intracellular metabolic enzymes contribute to the complex and unresolved phenomenon of multidrug resistance (MDR). Natural products as alternative medicine have great potential to discover new MDR inhibitors with diverse modes of action. In this study, we characterized several extracts of traditional Chinese medicine (TCM) plants (N = 16) for their interaction with ABC transporters, cytochrome P3A4 (CYP3A4), and glutathione-S-transferase (GST) activities and their cytotoxic effect on different cancer cell lines. Fallopia japonica (FJ) (Polygonaceae) shows potent inhibitory effect on CYP3A4 P-glycoprotein activity about 1.8-fold when compared to verapamil as positive control. FJ shows significant inhibitory effect (39.81%) compared with the known inhibitor ketoconazole and 100 μg/mL inhibited GST activity to 14 μmol/min/mL. FJ shows moderate cytotoxicity in human Caco-2, HepG-2, and HeLa cell lines; IC50 values were 630.98, 198.80, and 317.37 µg/mL, respectively. LC-ESI-MS were used to identify and quantify the most abundant compounds, emodin, polydatin, and resveratrol, in the most active extract of FJ. Here, we present the prospect of using Fallopia japonica as natural products to modulate the function of ABC drug transporters. We are conducting future study to evaluate the ability of the major active secondary metabolites of Fallopia japonica to modulate MDR and their impact in case of failure of chemotherapy.

Mechanism of Inhibition of CYP1A1 and Glutathione S-Transferase Activities in Fish Liver by Quercetin, Resveratrol, Naringenin, Hesperidin, and Rutin

Quercetin, resveratrol, naringenin, hesperidin, and rutin are phenolic compounds/flavonoids that may have roles in the reduction of cancer susceptibility. In this study, in vitro modulatory effects of them were studied on liver CYP1A1 associated 7-ethoxyresorufin-O-deethylase (EROD) activity and glutathione S-transferase (GST) activity of leaping mullet (Liza saliens). All of the phenolic compounds/flavonoids used exerted an inhibitory effect on both EROD and GST activities of fish. Quercetin, resveratrol, hesperidin, and rutin were found to inhibit EROD activity in a competitive manner; on the other hand, naringenin was found to inhibit EROD activity in a noncompetitive manner. Ki values of quercetin, resveratrol, naringenin, hesperidin, and rutin were calculated from Dixon plots as 0.12 μM, 0.67 μM, 2.63 μM, 18 μM and 0.1 mM, respectively. Resveratrol, quercetin, and hesperidin were found to inhibit GST activity in a competitive manner; on the other hand, rutin and naringenin were found to inhibit GST activity in a mixed-type manner. Ki values of resveratrol, quercetin, hesperidin, naringenin, and rutin were calculated from Dixon plots as 3.2 μM, 12.5 μM, 45 μM, 128 μM, and 150 μM, respectively. The results suggest that quercetin and resveratrol containing foods are effective in the prevention and treatment of cancer.

Inhibition of insect glutathione S-transferase (GST) by conifer extracts.

Insecticide synergists biochemically inhibit insect metabolic enzyme activity and are used both to increase the effectiveness of insecticides and as a diagnostic tool for resistance mechanisms. Considerable attention has been focused on identifying new synergists from phytochemicals with recognized biological activities, specifically enzyme inhibition. Jack pine (Pinus banksiana Lamb.), black spruce (Picea mariana (Mill.) BSP.), balsam fir (Abies balsamea (L.) Mill.), and tamarack larch (Larix laricina (Du Roi) Koch) have been used by native Canadians as traditional medicine, specifically for the anti-inflammatory and antioxidant properties based on enzyme inhibitory activity. To identify the potential allelochemicals with synergistic activity, ethanol crude extracts and methanol/water fractions were separated by Sephadex LH-20 chromatographic column and tested for in vitro glutathione S-transferase (GST) inhibition activity using insecticide-resistant Colorado potato beetle, Leptinotarsa decemlineata (Say) midgut and fat-body homogenate. The fractions showing similar activity were combined and analyzed by ultra pressure liquid chromatography-mass spectrometry. A lignan, (+)-lariciresinol 9'-p-coumarate, was identified from P. mariana cone extracts, and L.laricina and A. balsamea bark extracts. A flavonoid, taxifolin, was identified from P. mariana and P. banksiana cone extracts and L.laricina bark extracts. Both compounds inhibit GST activity with taxifolin showing greater activity compared to (+)-lariciresinol 9'-p-coumarate and the standard GST inhibitor, diethyl maleate. The results suggested that these compounds can be considered as potential new insecticide synergists.

3β-Acetyl tormentic acid reverts MRP1/ABCC1 mediated cancer resistance through modulation of intracellular levels of GSH and inhibition of GST activity

ABC transporter overexpression is an important mechanism of multidrug resistance (MDR) and one of the main obstacles to successful cancer treatment. As these proteins actively remove chemotherapeutics from the tumor cells, the pharmacological inhibition of their activity is a possible strategy to revert drug resistance. Moreover, the ability of MDR inhibitors to sensitize resistant cells to conventional drugs is important for their clinical use. Evidence has shown that the multidrug resistance protein 1 (MRP1/ABCC1) is a negative prognostic marker in patients with lung, gastric, or breast cancers or neuroblastoma. Previous data have shown that 3β-acetyl tormentic acid (3ATA) inhibits the transport activity of the protein MRP1/ABCC1. In this study, we evaluated the ability of 3ATA to sensitize an MDR cell line (GLC4/ADR), which overexpresses MRP1, and investigated the anti-MRP1 mechanisms activated by 3ATA. The results showed that 3ATA is able to reverse the resistance of the MDR cell line to doxorubicin and vincristine, two drugs that are commonly used in cancer chemotherapy. Regarding the sensitizing mechanism induced by 3ATA, this work shows that the triterpene does not modulate the expression of MRP1/ABCC1 but is able to reduce total intracellular glutathione (GSH) levels and decrease the activity of glutathione-s-transferase (GST), the enzyme responsible for the glutathione conjugation of xenobiotics. Together, these results show that 3ATA sensitizes the MDR cell line overexpressing MRP1/ABCC1 to antineoplastic drugs and that this effect is mediated by the modulation of intracellular levels of GSH and GST activity.

Individual and Mixture Toxicity of Commercial Formulations Containing Glyphosate, Metsulfuron-Methyl, Bispyribac-Sodium, and Picloram on Rhinella arenarum Tadpoles

We investigated the effects of four commercial formulations of herbicides (glyphosate [GLY], metsulfuron-methyl [MET], bispyribac-sodium [BIS], and picloram [PIC]) individually, and in three 50:50 mixtures (GLY–MET, GLY–BIS, GLY–PIC) on the common toad Rhinella arenarum (Anura: Bufonidae) tadpoles. Enzymatic parameters such as, glutathione S-transferase (GST), butyrylcholinesterase (BChE) and acetylcholinesterase (AChE) activities, as well as erythrocyte nuclear abnormalities (ENA) were studied. Interactions between herbicides in mixtures were evaluated and classified as additive, synergistic, or antagonistic. Toxicity results (48-h LC50) showed that PIC was the most toxic herbicide, followed by BIS, GLY, and MET, while GLY–PIC was the most toxic mixture, followed by GLY–BIS, and GLY–MET. All commercial herbicide formulations and their mixtures significantly inhibited BChE activity in exposed tadpoles. The AChE activity was also inhibited by all herbicides and their mixtures, except by GLY–BIS. The inhibition of GST activity was only significant for GLY, MET, PIC, and GLY–MET. A significant increase in the frequency of ENA was found for tadpoles exposed either to commercial herbicide formulations or to mixtures, except for GLY. All the mixtures showed synergism for BChE activity while for AChE only the GLY–MET and GLY–PIC mixtures acted synergistically. GLY–MET showed synergism for GST, whereas for ENA, the mixture GLY–BIS was antagonistic. This study with R. arenarum tadpoles demonstrates that the interactions between three of the most intensively used herbicides in soybean crops results in synergistic effects on mortality and neurotoxicity and synergistic or additive effects in genotoxicity.