Project description:Gene expression profiling reveals multiple tissue-specific functionality of GSH. We evaluated the effects of GSH against hydrogen peroxide (HP) on HepG2 cells. We performed an untargeted whole-genome transcriptome analysis to explore functionality of GSH.
Project description:Expression profiling of wild type E. coli K-12 strain MG1655 with exogenous glutathione (GSH) supplementation, sub-inhibitory ciprofloxacin concentration and GSH with inhibitory ciprofloxacin concentration. We used microarrays to detail the global gene expression program underlying ciprofloxacin exposure and glutathione mediated abrogation of ciprofloxacin's anti-bacterial action.
Project description:An excess of reactive oxygen species (ROS) can cause severe oxidative damage to cellular components in photosynthetic cells. Antioxidant systems, such as the ascorbate-glutathione cycle, regulate redox status in cells to guard against such damage. Dehydroascorbate reductase (DHAR, EC 1.8.5.1) catalyzes the glutathione-dependent reduction of oxidized ascorbate (dehydroascorbate) and contains a redox active site and glutathione binding-site. The DHAR gene is important in biological and abiotic stress responses involving reduction of the oxidative damage caused by ROS. In this study, a transgenic microalgae (TA) strain was constructed by cloning the Oryza sativa L. japonica DHAR (OsDHAR) gene controlled by an isopropyl β-D-1-thiogalactopyranoside (IPTG)-inducible promoter (Ptrc) into the Synechococcus elongatus PCC 7942 strain of cyanobacteria to study the functional activities of OsDHAR under oxidative stress caused by hydrogen peroxide exposure. OsDHAR expression increased the growth of S. elongatus PCC 7942 under oxidative stress by reducing the levels of hydroperoxides and malondialdehyde (MDA) and mitigating the loss of chlorophyll. DHAR and glutathione S-transferase activity were higher than in the wild-type (WT) strain. Additionally, overexpression of OsDHAR in S. elongatus PCC 7942 greatly increased the glutathione (GSH)/glutathione disulfide (GSSG) ratio compared to ascorbic acid (AsA)/dehydroascorbate (DHA) ratio in the presence or absence of hydrogen peroxide. These results strongly suggest that DHAR attenuates deleterious oxidative effects via the glutathione (GSH)-dependent antioxidant system in cyanobacterial cells. The expression of heterologous OsDHAR in S. elongatus PCC 7942 protected cells from oxidative damage through a GSH-dependent antioxidant system via GSH-dependent reactions at the redox active site and GSH binding site residues during oxidative stress.
Project description:Glutathione (GSH) is a tripeptide involved in controlling heavy metal movement in plants. Our previous study demonstrated that GSH, applied to plant roots site-specifically, inhibited Cd translocation from roots to shoots in oilseed rape plants (Brassica napus) cultured hydroponically. One of the factors of this inhibitory effect was due to activation of Cd efflux from root cells. To investigate the molecular mechanism triggered by root applied GSH in more detail, the Cd movement was monitored non-invasively using a positron-emitting tracer imaging system (PETIS). The Cd absorption and efflux process in roots were visualized successfully. The effects of GSH on Cd efflux from root cells were estimated by analyzing obtained imaging data. Another image analysis suggested that Cd return was activated by GSH, applied to roots, at the shoot base. Cutting the shoot base of oilseed rape plants significantly inhibited Cd efflux from root cells. These experimental results demonstrated the shoot base is playing important roles in distributing Cd in the plant bodies. Furthermore, DNA microarray analysis revealed that over 300 genes in the roots of oilseed rape plants responded to root applied GSH. Among them, transporter proteins, related to heavy metal movement in plants, and proteins related to changing the structure of cell walls were involved.
Project description:To comprehensively investigate the effects of glutathione on the gene expression, the microarray analysis was performed in the glutathione-fed wild-type Arabidopsis thaliana. Wild-type Arabidopsis (ecotype Columbia-0) were fed with 1 mM oxidized glutathione (GSSG) and 2 mM reduced glutathione (GSH) for comparison at equal nitrogen equivalents. To examine the effects of glutathione other than nitrogen at equal nitrogen equivalents, plants were fed with 3 mM NH4NO3. Plants grown by water were used as a control.
Project description:By microarray gene expression profiling, we investigated whether glutathione (GSH) has a signaling role in the response of RAW 264.7 cells to lipopolysaccharide (LPS). GSH was depleted by prior incubation with 120 µM buthionine sulfoximine (BSO) for 24 h, then cells were treated with 10 ng/ml LPS for 2 or 6 h. DNA microarray analysis was performed in triplicate samples in control and GSH-depleted cells, in the presence or absence of LPS.
Project description:Purpose: To better understand the effects of glutathione (GSH)-deficiency on lens homeostasis and cataractogenesis. Methods: The transcriptome of lens epithelia and fiber cells was obtained from C57BL/6 LEGSKO (lens GSH synthesis knockout) and buthionine sulfoximine (BSO)-treated LEGSKO mice and compared to C57BL/6 wild-type mice using RNA-Seq. Results: RNA-Seq results were in excellent agreement with qPCR (correlation coefficients between 0.87-0.94 and p<5E-6 for a subset of 36 mRNAs). Of the 24,415 transcripts mapped to the mouse genome, 441 genes showed significantly modulated expression. Pathway analysis indicated major changes in EMT signaling, visual cycle, small molecule biochemistry, and lipid metabolism. GSH-deficient lenses showed upregulation of genes relating to detoxification, including Aldh1a1, Aldh3a1 (aldehyde dehydrogenases), Mt1, Mt2 (metallothioneins), Ces1g (carboxylesterase), and Slc14a1 (urea transporter UT-B). These proteins share substrate specificity with GSH or glutathione-S-transferase and may protect GSH-deficient lenses. Genes associated with canonical EMT pathways, including Wnt10a, Egf, and Syk, showed upregulation in lens epithelia samples. Severely GSH-deficient lens epithelia showed a broad downregulation of vision-related genes (including Cryge, Crygf, and Rho). The BSO-treated LEGSKO lens epithelia transcriptome has significant correlation (r=0.63,P<0.005) to that of lens epithelia undergoing EMT. Conclusions: These results show that GSH depletion of the lens leads to expression of detoxifying genes and activation of EMT signaling, in addition to changes in transport systems and lipid homeostasis. These data give new insight into the adaptation and consequences of GSH-deficiency in the lens and suggest that supplementation of GSH or a precursor after cataract surgery could potentially reduce the incidence of EMT-mediated posterior subcapsular opacification.
Project description:Glutathione is a tripeptide involved in diverse aspects of plant metabolism. We investigated how the reduced form of glutathione, GSH, applied site-specifically to plants, affects zinc (Zn) distribution and behavior in oilseed rape plants (Brassica napus) cultured hydroponically. Foliar-applied GSH significantly increased the Zn content in shoots and the root-to-shoot Zn translocation ratio; furthermore, this treatment raised the Zn concentration in the cytosol of root cells and substantially enhanced Zn xylem loading. Notably, microarray analysis revealed that the gene encoding pectin methylesterase was upregulated in roots following foliar GSH treatment. We conclude that certain physiological signals triggered in response to foliar-applied GSH were transported via sieve tubes and functioned in root cells, which, in turn, increased Zn availability in roots by releasing Zn from their cell wall. Consequently, root-to-shoot translocation of Zn was activated and Zn accumulation in the shoot was markedly increased.
Project description:The essential thiol antioxidant, glutathione (GSH) is recruited into the nucleus of mammalian cells early in cell proliferation, suggesting a key role of the nuclear thiol pool in cell cycle regulation. However, the functions of nuclear GSH (GSHn) and its integration with the cytoplasmic GSH (GSHc) pools in whole cell redox homeostasis and signaling are unknown. Here we show that GSH is recruited into the nucleus early in cell proliferation in Arabidopsis thaliana, confirming the requirement for localization of GSH in the nucleus as a universal feature of cell cycle regulation. GSH accumulation in the nucleus was triggered by treatments that synchronize cells at G1/S as identified by flow cytometry and marker transcripts. Significant decreases in transcripts associated with oxidative signaling and stress tolerance occurred when GSH was localized in the nucleus. Increases in GSH1 and GSH2 transcripts accompanied the large increase in total cellular GSH observed during cell proliferation, but only GSH2 was differentially expressed in cells with high GSHn relative to those with an even intracellular distribution of GSH. Of the 7 Bcl-2 associated (BAG) genes in A. thaliana, only the nuclear-localized BAG 6 was differentially expressed in cells with high GSHn compared to GSHc. We conclude that GSHn is associated with decreased oxidative signaling and stress responses and that whole cell redox homeostasis is restored as the cell cycle progresses by enhanced GSH synthesis and accumulation in the cytoplasm. Arabidopsis cells were harvested at points during cell proliferation where GSH was localized either in the nucleus (GSHn) or where GSH was distributed throughout the cytoplasm (GSHc) for RNA extraction and hybridization on Affymetrix microarrays. We selected three stages where the GSH was into the nucleus and three stages where the GSH was distributed throughout the cells.
Project description:Background: Autologous fat grafting is hampered by unpredictable graft survival, which is potentially regulated by ferroptosis. Glutathione (GSH), a powerful antioxidant used in tissue preservation, has ferroptosis-regulating activity; however, its effects on fat grafts are unclear. This study investigated the effects and mechanisms of GSH in fat graft survival. Methods: Human lipoaspirates were transplanted subcutaneously into the backs of normal saline-treated (control) or GSH-treated nude mice. Graft survival was evaluated by magnetic resonance imaging and histology. RNA sequencing was performed to identify differentially expressed genes and enriched pathways. GSH activity was evaluated in vitro using an oxygen and glucose deprivation (OGD) model of adipose-derived stem cells. Results: Compared with control group, GSH induced better outcomes, including superior graft retention, appearance, and histological structures. RNA sequencing suggested enhanced negative regulation of ferroptosis in the GSH-treated grafts, which showed reduced lipid peroxides, better mitochondrial ultrastructure, and SLC7A11/GPX4 axis activation. In vitro, OGD-induced ferroptosis was ameliorated by GSH, which restored cell proliferation, reduced oxidative stress, and upregulated ferroptosis defense factors. Conclusions: Our study confirms that ferroptosis participates in regulating fat graft survival and that GSH exerts a protective effect by inhibiting ferroptosis. GSH-assisted lipotransfer is a promising therapeutic strategy for future clinical application.