Narrow Results

This study investigated the effects of various concentrations and incubation time intervals of CDA-II (cell differentiation agent: a preparation of human urine) on cell viability, lipid peroxidation and glutathione and its related enzyme activities in rat hepatocytes. Based on the results of lactate dehydrogenase leakage and microscopic examination, treatment with CDA-II significantly elevated the viability of hepatocytes. The level of lipid peroxidation of cells treated with 2.5 or 5 mg/ml CDA-II was lower than the control after 4, 8 and 24 hours of incubation. Intracellular glutathione (GSH) content of cells treated with 0.25–5 mg/ml CDA-II was significantly higher than controls after 8 and 24 hours of incubation. These phenomena are beneficial to the antioxidant capabilities of hepatocytes. Furthermore, treatment of hepatocytes with CDA-II has no effect on the activities of GSH peroxidase, GSH reductase and GSH S-transferase. In conclusion, adequate concentration of CDA-II could enhance the viability and antioxidative capability of hepatocytes.

The objective of this study was to investigate the effects of various concentrations and incubation times of water extract of clam (WEC) on glutathione, its antioxidant and the detoxification defense systems in normal and CCl₄-induced oxidative damaged primary rat hepatocytes. This study showed that when the hepatocytes were treated with WEC(0.14 ~ 1.68 mg/ml), the intracellular glutathione (GSH) levels, GSH/GSSG ratio, and the activities of GSH-related enzymes (GPx, GRd, and GST) were higher than those in the control at 24 or 48 hour treatments. However, the lactate dehydrogenase (LDH) leakage and microscopic observations did not differ from those of the control. Yet, when the hepatocytes were pretreated with various concentrations of WEC for 24 hours and then exposed to 5 mM carbon tetrachloride (CCl₄) for 1 hour, at concentrations of WEC between 0.42 ~ 1.68 mg/ml, the viabilities, intracellular GSH level, and activities of GST and GPx were significantly increased compared to those of the CCl₄-treated control group (p < 0.05). In conclusion, WEC could improve the viability and the capabilities of detoxification and antioxidation in hepatocytes by increasing the GSH level and the activities of GSH-related enzymes.

(1′S)-acetoxychavicol acetate ((S)-ACA) exhibits chemopreventive effects on chemically induced tumor formation. It has been shown that ACA inhibited the development of azoxymethane-induced colon carcinogenesis through its suppression of cell proliferation in the colonic mucosa and its induction of glutathione S-transferase and quinone oxidoreductase 1 in vivo. In this study, we investigated how ACA induced these enzymes by using rat intestine epithelial cells (IEC6) in vitro. ACA induced glutathione S-transferase (GST) and NAD (P)H: quinone oxidoreductase 1 (NQO1) activities, increased intracellular glutathione (GSH) level, and upregulated intranuclear Nrf2 and cytosolic p21. It suggested that activation of phase II enzymes via Nrf2 associated with p21 is one of possible mechanisms of ACA to prevent advance of carcinogenesis.

The activation of cobalamin requires a reduction from cobalamin(III) to cobalamin(II). The reduction by glutathione and dithiothreitol was followed using visible spectroscopy and electron paramagnetic resonance. In addition the oxidation of glutathione was monitored. Glutathione first reacts with oxidized cobalamin(III). The binding of a second glutathione required for the reduction to cobalamin(II) is presumably located in the dimethyl benzimidazole ribonucleotide ligand cavity. The reduction of cobalamin(III) by dithiothreitol, which contains two thiols, is much faster even though no stable cobalamin(III) complex is formed. The reduction, by both thiol reagents, results in the formation of thiyl radicals, some of which are released to form oxidized thiol products and some of which remain associated with the reduced cobalamin. In the reduced state the intrinsic lower affinity for the benzimidazole base, coupled with a trans effect from the initial glutathione bound to the β-axial site and a possible lowering of the pH results in an equilibrium between base-on and base-off complexes. The dissociation of the base facilitates a closer approach of the thiyl radical to the cobalamin(II) α-axial site resulting in a complex with ferromagnetic exchange coupling between the metal ion and the thiyl radical. This is a unique example of "internal spin trapping" of a thiyl radical formed during reduction. The finding that the reduction involves a peripheral site and that thiyl radicals produced during the reduction remain associated with the reduced cobalamin provide important new insights into our understanding of the formation and function of cobalamin enzymes.

Three derivatives of chlorin e6 (1–3) were synthesized by introduction of maleimide, cysteine and glutathione at C-13 carboxyl of the chlorin scaffold. The evaluation of their PDT effects showed that compound 1, the derivative with a maleimide group, exhibited more potent photocytotoxicity against HepG2 cells (IC${}_{50}$ 3.2 $\mu$M) than 2 (IC${}_{50}$ 6.7 $\mu$M) and 3 (IC${}_{50}$ 10.2 $\mu$M), although the cellular uptake of 1 was slightly lower than that of 2 and 3. The high PDT effect of 1 was found to be in agreement with the high level of intracellular singlet oxygen. Further investigation of the mechanism revealed that 1 can significantly lower the GSH level in HepG2 cells due to the addiction reaction of maleimide and GSH, thus resulting in the reduction of ROS scavenging and the enhancement of cellular oxidative stress. This approach to improve PDT effects of photosensitizers by means of interfering with the cellular redox system and enhancing cellular oxidative stress offers a new strategy for development of photosensitizers in cancer therapy.

Here, we suggest a new method of increasing the reactivity of cyanocobalamin (CNCbl) and its meso-chlorinated and brominated derivatives (CNCbl-Cl,Br) toward glutathione (GSH) that facilitates their reductive decyanation. In particular, we found that introducing a $c$-lactone ring to CNCbl or its halogenated versions substantially accelerates their reduction by GSH to a Co(II)-species. The highest rate of the reaction was observed in the case of CNCbl-Cl $c$-lactone. The mechanism of CNCbl $c$-lactone reduction by GSH involves the formation of intermediate complexes with GSH, which are further transformed into unstable Co(III)-thiolato species with detached 5,6-dimethylbenzimidazole (DMBI) decomposing to Co(II)-species and thiyl radical. In the case of the reaction involving CNCbl-Cl $c$-lactone, a parallel reduction pathway involves direct DMBI substitution by GSH. Besides the formation of Co(II)-species, the reduction of lactone ring occurs in the course of reactions of CNCbl $c$-lactones with GSH, giving species possessing lower reactivity toward GSH than initial $c$-lactones.

Hispolon is an active ingredient contained in the medicinal mushroom Phellinus linteus (PL) that is used in traditional Chinese medicine for various remedies, including lymphatic diseases. Previous studies reported that hispolon exhibited anti-inflammatory activities and suppressed mitogen-induced proliferation of splenic lymphocytes. It remains unclear if hispolon influences antigen-specific immunity. The present study investigated the effects of hispolon on cytokine production by antigen-activated T cells. Ovalbumin (OVA)-primed splenocytes were exposed to hispolon, followed by restimulation with OVA. Cell viability was determined by the AlamarBlue^® assay and T cell cytokine production was measured by enzyme linked immunosorbent assay (ELISA). The splenocyte viability and the production of interleukin (IL)-4 were unaffected, whereas the production of interferon (IFN)-γ was significantly suppressed by treatment with hispolon (1–5 μM) in a concentration-dependent manner. The suppressive effect of hispolon on the production of IFN-γ was attenuated in the presence of thiol antioxidants, including N-acetyl-_L-cysteine (NAC) and glutathione, whereas the non-thiol antioxidant pyruvate was ineffective. Taken together, these results demonstrated that hispolon induced a differential effect on antigen-induced cytokine production by T cells, in which the T helper 1 (Th1) cytokine IFN-γ was sensitive, whereas the Th2 cytokine IL-4 was unaffected. In addition, the suppressive effect of hispolon on IFN-γ production was associated with the diminishment of intracellular glutathione.

Herein, we present the one-pot synthesis of carbon quantum dots (CQDs) using glutathione and used for the optical detection of hydrogen peroxide (H₂O₂). Unlike as reported in other literatures, the detection of H₂O₂ is free of using any additional chromogenic agents or metal ions. The CQDs are synthesized using glutathione by one-pot hydrothermal at 180^∘C. The as-prepared CQDs exhibit good fluorescence response toward H₂O₂ without involving extra chromogenic substances. The addition of 300$\mu$M H₂O₂ to CQDs induces the quenching of fluorescence, besides the yellow becomes colorless. The decrease in the fluorescence is attributed to the polymeric structure of CQDs disbanded by H₂O₂. In addition, the practical application of the present method was verified by determining H₂O₂ in human urine samples.

Photoelectrochemical (PEC) sensor is an important type of biosensor widely used in glutathione (GSH) sensing. The PEC properties of the photoanode present in the sensor are critical to its sensing performance. Zinc oxide (ZnO) is an excellent semiconductor with a suitable band gap and light absorption ability for photoanode applications. Meanwhile, the interfacial layer is also important in the separation and transportation process of the excitons. In this work, high-quality ZnO nanorods were grown on the indium tin oxide (ITO) substrates. An interfacial layer consisting of reduced graphene oxide (RGO) or MXene (a two-dimensional transition metal carbide)-derived TiO₂ was introduced. Our results show that the introduction of the RGO/TiO₂ hybrid interfacial layer can promote both the high-quality growth of ZnO nanorods and also provides suitable band gap grading for efficient excitons separation and transportation. The GSH sensing performance of the PEC sensor based on the ZnO nanorods grown on the RGO/TiO₂ hybrid layer-coated ITO photoanode can dramatically improve the photocurrent strength and linearity.

To enhance the poor sensing capabilities of traditional p-type metal oxide semiconductor sensors, a novel gas sensor was designed with Co₃O₄/CuO heterostructure modified by glutathione (GSH)-reduced gold (Au), which was characterized by different analyses. The experiments showed that the 0.5wt.% Au–Co₃O₄/CuO gas sensor exhibited excellent sensing performance with a response value of up to 28.3 for 3ppm NO₂ at a relatively low operating temperature ($100^{\circ }$C). Besides, it demonstrated a linear relationship between detection concentration and response value, displayed good selectivity for NO₂, showcased stability and exhibited repeatability. Moreover, the sensor revealed the ability to resist humidity at the optimal temperature. Finally, we proposed a gas sensing mechanism to explain the enhanced NO₂ detection performance of the sensor, attributed to the heterojunction structure between Co₃O₄/CuO and the catalytic properties of Au. This led to an increase in adsorbed oxygen, as confirmed by XPS characterization. Therefore, the 0.5wt.% Au–Co₃O₄/CuO ternary nanocomposite gas sensor is deemed to be a highly effective sensing material that can be utilized to detect NO₂ gas in real-life applications.

Surface-enhanced Raman scattering (SERS) has been widely studied and applied for over three decades. However, reliable SERS detection of molecules with low polarizability is still suffering from poor sensitivity and reproducibility. In this paper, we have reported a new strategy for performing quantitative SERS detection of Raman insensitive Glutathione (GSH), based on GSH-induced replacement of a highly Raman sensitive four-mercaptopyridine (MP) adsorbed on the surface of four-aminothiophenol (ATP) embedded Au-core/Ag-shell particles. This replacement led to a strong decrease of the MP SERS signal, which was used to determine the concentration of GSH. The adoption of GSH-induced Raman probe replacement leads to high sensitivity, while the use of internal reference method provides an improved accuracy of the GSH quantification.

The following sections are included:

• Summary

• Methods Developed to Study Protein Thiols
  • Selective Radioactive Protein Thiol Labeling Reagent N-iodoacetyl-[¹²⁵I]-3-iodotyrosine (IAIT)
  • Arsenical-Based Affinity Chromatography of Vicinal Thiol Proteins
  • Thiol/Disulfide Exchange to Purify Monothiol Proteins
  • Determination of the Redox State of Thiol Proteins in Intact Cells

• Nature and Redox State of Thiol Proteins in Cell Extracts
  • The Majority of Cell Proteins are Present in the Reduced State
  • The Majority of the Cell Thiol Proteins Contain Surface-Localized Vicinal Thiols
  • Properties of Thiol Proteins Oxidized by Reaction with Active Disulfides
  • NADPH-Dependent Protein Disulfide Reductase Activity Depends on the Mixed Disulfide Formed by Thioredoxin
  • Implications of the Studies in vitro with Oxidized Cell Extracts

• Redox Regulation during Growth Initiation⁹
  • Growth Factors Induce a Calcium-Dependent Transient Oxidation of TXN and Other Cell Proteins⁹
  • Calcium Inhibits the in vitro Activity of Thioredoxin Reductase⁹
  • Implications of the Transient Oxidation of Thioredoxin and Other Cell Dithiol Proteins

• Concluding Remarks

• Acknowledgments

• References

The following sections are included:

• Introduction

• The Thioredoxin System

• The Glutaredoxin System

• Redox Regulation by Thioredoxin and Glutaredoxin

• Effects of Oxidants and Antioxidants

• Future Perspectives

• Acknowledgments

• References

The diuron is an herbicide used on autumn-winter crops, due to its ability to block the chloroplast electron chain at level of photosystem II. Furthermore, diuron can also exert toxic effects on heterotrophic beings, blocking the respiratory chain and ROS generating. Despite the progressive suppression of diuron application by Directive 200/60/CE, groundwater contamination will still persist. Therefore the main purpose of this work was to evaluate the effect of this phenylurea on S. cerevisiae at proliferative phase. The results show a significant decrease of GSH/GSSG ratio, in cells exposed to 50 μM diuron. Although GR activity does not changes significantly in any assay of this study, it was observed a significantly decrease of G6PD and GPx enzyme activities, in cells grown in 50 and 75 μM diuron. The decline of NADPH availability probably blocks the glutathione cycle, generating oxidative stress. In addition, the CAT T activity presents also a significant increase in cells grown in 50 μM diuron, as well as an increase of cytoplasm ROS, MDA level and CAT A activity in cultures with 50 and 75 μM diuron, that suggests a key role of peroxisomal lipid oxidation in diuron response which cause cell death by an active process.

Autism spectrum disorders (ASDs) are complex multifactorial disorders involving various genetic vulnerabilities interacting with environmental factors. Although the cause of autism is elusive, numerous studies indicate that oxidative stress and mitochondrial dysfunction are involved in the pathophysiology of ASDs. Several studies have reported increased levels of oxidative damage markers of lipid peroxidation, protein and DNA oxidation as well as impaired antioxidant defense in the plasma, urine and/or brain tissues from autistic subjects. We and other groups have also reported decreased glutathione (GSH) level and GSH/GSSG (oxidized glutathione) ratio, and increased GSSG in the blood and brain samples of autistic subjects, suggesting glutathione redox imbalance in autism. The activities of enzymes involved in GSH homeostasis i.e. glutathione peroxidase (GPx), glutathione-S-transferase (GST) and glutamate cysteine ligase (GCL) are impaired in autism, and association of polymorphisms of GSH, GPx or GST genes with autism has been reported. Mitochondria are not only the source of free radicals, but they are also the target of oxidative damage. Electron transport chain (ETC) in mitochondria is essential for ATP generation. Defects in energy metabolism, protein expression and activities of ETC complexes as well as in mitochondrial DNA have been reported in the blood, muscle biopsy and brain samples from autism. All these findings suggest that oxidative stress and mitochondrial dysfunction play critical roles in the pathophysiology of autism.

Despite decades of research, effective medical treatments for autism spectrum disorders are very limited. In fact, the idea that autistic children can be successfully treated to achieve optimal outcomes has only recently become more broadly accepted. Recent studies have demonstrated an association between autism and several metabolic abnormalities, including disorders of mitochondrial, folate, redox, glutathione and tetrahydrobiopterin metabolism. Many of these metabolic abnormalities have the potential to be amenable to novel treatments, thereby opening the door for improvements and potential recovery from the disabilities that define and are associated with autism. In fact, several preliminary clinical studies have suggested positive effects from treatment of these metabolic abnormalities. This chapter reviews the metabolic abnormalities that are potentially amenable to treatment, and the evidence for the effectiveness of treatments of these physiological abnormalities in autism. Although preliminary, this evidence paves the way for achieving optimal outcomes in children with autism.