Sulforaphane prevents diabetes-induced hepatic ferroptosis by activating Nrf2 signaling axis.

Recently, we characterized the ferroptotic phenotype in the liver of diabetic mice and revealed nuclear factor (erythroid-derived-2)-related factor 2 (Nrf2) inactivation as an integral part of hepatic injury. Here, we aim to investigate whether sulforaphane, an Nrf2 activator and antioxidant, prevents diabetes-induced hepatic ferroptosis and the mechanisms involved. Male C57BL/6 mice were divided into four groups: control (vehicle-treated), diabetic (streptozotocin-induced; 40 mg/kg, from Days 1 to 5), diabetic sulforaphane-treated (2.5 mg/kg from Days 1 to 42) and non-diabetic sulforaphane-treated group (2.5 mg/kg from Days 1 to 42). Results showed that diabetes-induced inactivation of Nrf2 and decreased expression of its downstream antiferroptotic molecules critical for antioxidative defense (catalase, superoxide dismutases, thioredoxin reductase), iron metabolism (ferritin heavy chain (FTH1), ferroportin 1), glutathione (GSH) synthesis (cystine-glutamate antiporter system, cystathionase, glutamate-cysteine ligase catalitic subunit, glutamate-cysteine ligase modifier subunit, glutathione synthetase), and GSH recycling - glutathione reductase (GR) were reversed/increased by sulforaphane treatment. In addition, we found that the ferroptotic phenotype in diabetic liver is associated with increased ferritinophagy and decreased FTH1 immunopositivity. The antiferroptotic effect of sulforaphane was further evidenced through the increased level of GSH, decreased accumulation of labile iron and lipid peroxides (4-hydroxy-2-nonenal, lipofuscin), decreased ferritinophagy and liver damage (decreased fibrosis, alanine aminotransferase, and aspartate aminotransferase). Finally, diabetes-induced increase in serum glucose and triglyceride level was significantly reduced by sulforaphane. Regardless of the fact that this study is limited by the use of one model of experimentally induced diabetes, the results obtained demonstrate for the first time that sulforaphane prevents diabetes-induced hepatic ferroptosis in vivo through the activation of Nrf2 signaling pathways. This nominates sulforaphane as a promising phytopharmaceutical for the prevention/alleviation of ferroptosis in diabetes-related pathologies.

The Key Role of GSH in Keeping the Redox Balance in Mammalian Cells: Mechanisms and Significance of GSH in Detoxification via Formation of Conjugates.

Glutathione (GSH) is a ubiquitous tripeptide that is biosynthesized in situ at high concentrations (1-5 mM) and involved in the regulation of cellular homeostasis via multiple mechanisms. The main known action of GSH is its antioxidant capacity, which aids in maintaining the redox cycle of cells. To this end, GSH peroxidases contribute to the scavenging of various forms of ROS and RNS. A generally underestimated mechanism of action of GSH is its direct nucleophilic interaction with electrophilic compounds yielding thioether GSH S-conjugates. Many compounds, including xenobiotics (such as NAPQI, simvastatin, cisplatin, and barbital) and intrinsic compounds (such as menadione, leukotrienes, prostaglandins, and dopamine), form covalent adducts with GSH leading mainly to their detoxification. In the present article, we wish to present the key role and significance of GSH in cellular redox biology. This includes an update on the formation of GSH-S conjugates or GSH adducts with emphasis given to the mechanism of reaction, the dependence on GST (GSH S-transferase), where this conjugation occurs in tissues, and its significance. The uncovering of the GSH adducts' formation enhances our knowledge of the human metabolome. GSH-hematin adducts were recently shown to have been formed spontaneously in multiples isomers at hemolysates, leading to structural destabilization of the endogenous toxin, hematin (free heme), which is derived from the released hemoglobin. Moreover, hemin (the form of oxidized heme) has been found to act through the Kelch-like ECH associated protein 1 (Keap1)-nuclear factor erythroid 2-related factor-2 (Nrf2) signaling pathway as an epigenetic modulator of GSH metabolism. Last but not least, the implications of the genetic defects in GSH metabolism, recorded in hemolytic syndromes, cancer and other pathologies, are presented and discussed under the framework of conceptualizing that GSH S-conjugates could be regarded as signatures of the cellular metabolism in the diseased state.

MTX-211 Inhibits GSH Synthesis through Keap1/NRF2/GCLM Axis and Exerts Antitumor Effects in Bladder Cancer.

Globally, bladder cancer (BLCA) is still the leading cause of death in patients with tumors. The function and underlying mechanism of MTX-211, an EFGR and PI3K kinase inhibitor, have not been elucidated. This study examined the function of MTX-211 in BLCA cells using in vitro and in vivo assays. RNA sequencing, quantitative real-time polymerase chain reaction, Western blotting, co-immunoprecipitation, and immunofluorescence were performed to elucidate the underlying mechanism. Our observations revealed that MTX-211 has a time- and concentration-dependent inhibitory effect on bladder cancer cell proliferation. Flow cytometry analysis showed that cell apoptosis and G0/G1 cell cycle arrest were significantly induced by MTX-211. MTX-211 inhibited intracellular glutathione (GSH) metabolism, leading to a decrease in GSH levels and an increase in reactive oxygen species. GSH supplementation partly reversed the inhibitory effects of MTX-211. Further experiments verified that MTX-211 promoted NFR2 protein ubiquitinated degradation via facilitating the binding of Keap1 and NRF2, subsequently resulting in the downregulated expression of GCLM, which plays a vital role in GSH synthesis. This study provided evidence that MTX-211 effectively inhibited BLCA cell proliferation via depleting GSH levels through Keap1/NRF2/GCLM signaling pathway. Thus, MTX-211 could be a promising therapeutic agent for cancer.

Azithromycin ameliorated cigarette smoke-induced airway epithelial barrier dysfunction by activating Nrf2/GCL/GSH signaling pathway.

BACKGROUND: Airway epithelium is the first barrier against environmental insults, and epithelial barrier dysfunction caused by cigarette smoke (CS) is particularly relevant to chronic obstructive pulmonary disease (COPD) progression. Our study was to determine whether Azithromycin (AZI) ameliorates CS-induced airway epithelial barrier dysfunction and the underlying mechanisms. METHODS: Primary bronchial epithelial cells (PBECs), human bronchial epithelial cells (HBECs), Sprague Dawley rats and nuclear factor erythroid 2-related factor 2 (Nrf2)-/- mice were pretreated with AZI and subsequently exposed to CS. Transepithelial electronic resistance (TEER), junction proteins as well as pro-inflammatory cytokines and apoptosis markers were examined to assess epithelial barrier dysfunction. Metabolomics study was applied to explore the underlying mechanism of AZI. RESULTS: CS-induced TEER decline and intercellular junction destruction, accompanied with inflammatory response and cell apoptosis in PBECs were restored by AZI dose-dependently, which were also observed in CS-exposed rats. Mechanistically, GSH metabolism pathway was identified as the top differentially impacted pathway and AZI treatment upregulated the activities of glutamate cysteine ligase (GCL) and the contents of metabolites in GSH metabolic pathway. Furthermore, AZI apparently reversed CS-induced Nrf2 suppression, and similar effects on airway epithelial barrier dysfunction were also found for Nrf2 agonist tert-butylhydroquinone and vitamin C. Finally, deletion of Nrf2 in both HBECs and C57BL/6N mice aggravated CS-induced GSH metabolism imbalance to disrupt airway epithelial barrier and partially deprived the effects of AZI. CONCLUSION: These findings suggest that the clinical benefits of AZI for COPD management are related with the protection of CS-induced airway epithelial barrier dysfunction via activating Nrf2/GCL/GSH pathway, providing potential therapeutic strategies for COPD.

Sustainable and efficient method utilizing N-acetyl-L-cysteine for complete and enhanced ochratoxin A clearance by antagonistic yeast.

With the increasing global climate change, ochratoxin A (OTA) pollution in food and environment has become a serious and potential risk element threatening food safety and human health. Biodegradation of mycotoxin is an eco-friendly and efficient control strategy. Still, research works are warranted to develop low-cost, efficient, and sustainable approaches to enhance the mycotoxin degradation efficiency of microorganisms. In this study, the activities of N-acetyl-L-cysteine (NAC) against OTA toxicity were evidenced, and its positive effects on the OTA degradation efficiency of antagonistic yeast, Cryptococcus podzolicus Y3 were verified. Co-culturing C. podzolicus Y3 with 10 mM NAC improved 100% and 92.6% OTA degradation rate into ochratoxin α (OTα) at 1 d and 2 d. The excellent promotion role of NAC on OTA degradation was observed even at low temperatures and alkaline conditions. C. podzolicus Y3 treated with OTA or OTA+NAC promoted reduced glutathione (GSH) accumulation. GSS and GSR genes were highly expressed after OTA and OTA+NAC treatment, contributing to GSH accumulation. In the early stages of NAC treatment, yeast viability and cell membrane were reduced, but the antioxidant property of NAC prevented lipid peroxidation. Our finding provides a sustainable and efficient new strategy to improve mycotoxin degradation by antagonistic yeasts, which could be applied to mycotoxin clearance.

Physio-Biochemical and Transcriptomic Features of Arbuscular Mycorrhizal Fungi Relieving Cadmium Stress in Wheat.

Arbuscular mycorrhizal fungi (AMF) can improve plant cadmium (Cd) tolerance, but the tolerance mechanism in wheat is not fully understood. This study aimed to examine the physiological properties and transcriptome changes in wheat inoculated with or without Glomus mosseae (GM) under Cd stress (0, 5, and 10 mg·kg-1 CdCl2) to understand its role in wheat Cd tolerance. The results showed that the Cd content in shoots decreased while the Cd accumulation in roots increased under AMF symbiosis compared to the non-inoculation group and that AMF significantly promoted the growth of wheat seedlings and reduced Cd-induced oxidative damage. This alleviative effect of AMF on wheat under Cd stress was mainly attributed to the fact that AMF accelerated the ascorbate-glutathione (AsA-GSH) cycle, promoted the production of GSH and metallothionein (MTs), improved the degradation of methylglyoxal (MG), and induced GRSP (glomalin-related soil protein) secretion. Furthermore, a comparative analysis of the transcriptomes of the symbiotic group and the non-symbiotic group revealed multiple differentially expressed genes (DEGs) in the 'metal ion transport', 'glutathione metabolism', 'cysteine and methionine metabolism', and 'plant hormone signal transduction' terms. The expression changes of these DEGs were basically consistent with the changes in physio-biochemical characteristics. Overall, AMF alleviated Cd stress in wheat mainly by promoting immobilization and sequestration of Cd, reducing ROS production and accelerating their scavenging, in which the rapid metabolism of GSH may play an important role.

Advancement in understanding the role of ferroptosis in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic, systemic disease of unknown etiology. The primary manifestation of RA is inflammatory synovitis, which eventually leads to deformity and functional loss. Ferroptosis is a non-apoptosis form of cell death that depends on intracellular iron accumulation. This leads to an increase in reactive oxygen species (ROS) induced-lipid peroxidation. The underlying mechanisms of ferroptosis are System Xc- and Glutathione metabolism, regulation of glutathione peroxidase 4 activity, and ROS generation. Recent studies have shown an association between the pathogenesis of RA and ferroptosis, suggesting the involvement of ferroptosis in the onset and progression of RA. In this review, we have focused on the mechanism of ferroptosis and its association with RA pathogenesis. Further, we discuss the status of therapeutics targeting ferroptosis in the treatment of patients with RA. Targeting ferroptosis could be a potential therapeutic approach for RA treatment.

Unraveling Cadmium Toxicity in Trifolium repens L. Seedling: Insight into Regulatory Mechanisms Using Comparative Transcriptomics Combined with Physiological Analyses.

Trifolium repens (T. repens) can accumulate significant amounts of heavy metal ions, and has strong adaptability to wide environmental conditions, and relatively large biomass, which is considered a potential plant for phytoremediation. However, the molecular mechanisms of T. repens involved in Cd tolerance have not yet been studied in detail. This study was conducted to examine the integrative responses of T. repens exposed to a high-level CdCl2 by investigating the physiological and transcriptomic analyses. The results suggested that T. repens seedlings had a high degree of tolerance to Cd treatment. The roots accumulated higher Cd concentration than leaves and were mainly distributed in the cell wall. The content of MDA, soluble protein, the relative electrolyte leakage, and three antioxidant enzymes (POD, SOD, and APX) was increased with the Cd treatment time increasing, but the CAT enzymes contents were decreased in roots. Furthermore, the transcriptome analysis demonstrated that the differentially expressed genes (DEGs) mainly enriched in the glutathione (GSH) metabolism pathway and the phenylpropanoid biosynthesis in the roots. Overexpressed genes in the lignin biosynthesis in the roots might improve Cd accumulation in cell walls. Moreover, the DEGs were also enriched in photosynthesis in the leaves, transferase activity, oxidoreductase activity, and ABA signal transduction, which might also play roles in reducing Cd toxicity in the plants. All the above, clearly suggest that T. repens employ several different mechanisms to protect itself against Cd stress, while the cell wall biosynthesis and GSH metabolism could be considered the most important specific mechanisms for Cd retention in the roots of T. repens.

Regulation of ferroptosis by noncoding RNAs: a novel promise treatment in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is a highly prevalent tumor that requires extensive research. Ferroptosis is a unique cell death modality driven by iron-dependent phospholipid peroxidation manifested as an accumulation of lipid-reactive oxygen species. With further understanding of noncoding RNAs (ncRNAs), numerous studies have demonstrated an important regulatory role of ncRNAs in ESCC through ferroptosis, including microRNAs, long ncRNAs, and circular RNAs. These ncRNAs influence the expression of the target gene to regulate ESCC progression by involving the ferroptosis signaling pathway. However, the specific regulatory mechanism of ncRNAs on ferroptosis in ESCC remains largely unknown. This review summarized the current knowledge on the relation between ferroptosis regulators, such as glutathione synthesis/metabolism, Keap1/Nfr2, and p53, by ncRNAs and ESCC. This review also proposed the possible therapeutic approaches for ncRNAs targeting ferroptosis in ESCC. This is the latest and most effective summary of recent research achievements of ncRNAs on ferroptosis in ESCC. These ncRNAs based on ferroptosis merit further investigation in preclinical research of ESCC.

Niujiaodihuang Detoxify Decoction inhibits ferroptosis by enhancing glutathione synthesis in acute liver failure models.

ETHNOPHARMACOLOGICAL RELEVANCE: Niujiaodihuang Detoxify Decoction (NDD) is an integrated traditional Chinese medicine prescription that has been used as a therapeutic agent for the treatment of acute liver failure (ALF). However, the mechanisms underlying its action remain unclear. AIM OF THE STUDY: To determine the protective effect of NDD on D-galactosamine/lipopolysaccharide (D-GalN/LPS)-induced ALF and explore the underlying mechanisms. MATERIALS AND METHODS: We characterized the NDD fingerprint by HPLC and established D-GalN/LPS-induced ALF models in Sprague-Dawley rats and LO2 cells. Next, we measured the protective and antiferroptotic effects of NDD in vivo and in vitro. To further investigate the molecular mechanisms underlying the effects of NDD, we performed metabolomic analysis of the liver tissue using LC-MS/MS. RESULTS: Results of serum biochemical analysis, liver histopathology, and cell viability showed that NDD effectively relieved the liver injury. It reduced the accumulation of labile iron and alleviated lipid peroxidation by enhancing GPX4 activity. The mitochondrial morphology indicated that NDD exerted its hepatoprotective effect through an antiferroptotic activity. Metabolomic analysis showed that NDD treatment increased the levels of cysteine, decreased those of glutamate, and ameliorated the D-GalN/LPS-induced reduction in the levels of glutathione (GSH). The results for intracellular levels of reduced (GSH) and oxidized (GSSG) glutathione were consistent with those of metabolomic analysis. CONCLUSION: Our findings indicate that NDD exerts hepatoprotective activity by evoking the reprogramming of GSH metabolism, and thereby, inhibiting ferroptosis.

The Glutathione Metabolite γ-Glutamyl-Glutamate Partially Activates Glutamate NMDA Receptors in Central Neurons With Higher Efficacy for GluN2B-Containing Receptors.

Gamma-L-glutamyl-L-glutamate (γ-Glu-Glu) was synthetized and further characterized for its activity on cultured neurons. We observed that γ-Glu-Glu elicited excitatory effects on neurons likely by activating mainly the N-methyl-D-aspartate (NMDA) receptors. These effects were dependent on the integrity of synaptic transmission as they were blocked by tetrodotoxin (TTX). We next evaluated its activity on NMDA receptors by testing it on cells expressing these receptors. We observed that γ-Glu-Glu partially activated NMDA receptors and exhibited better efficacy for NMDA receptors containing the GluN2B subunit. Moreover, at low concentration, γ-Glu-Glu potentiated the responses of glutamate on NMDA receptors. Finally, the endogenous production of γ-Glu-Glu was measured by LC-MS on the extracellular medium of C6 rat astroglioma cells. We found that extracellular γ-Glu-Glu concentration was, to some extent, directly linked to GSH metabolism as γ-Glu-Glu can be a by-product of glutathione (GSH) breakdown after γ-glutamyl transferase action. Therefore, γ-Glu-Glu could exert excitatory effects by activating neuronal NMDA receptors when GSH production is enhanced.

The effect of toxic components on metabolomic response of male SD rats exposed to fine particulate matter.

PM2.5 pollution was associated with numerous adverse health effects. However, PM2.5 induced toxic effects and the relationships with toxic components remain largely unknown. To evaluate the metabolic toxicity of PM2.5 at environmentally relevant doses, investigate the seasonal variation of PM2.5 induced toxicity and the relationship with toxic components, a combination of general pathophysiological tests and metabolomics analysis was conducted in this study to explore the response of SD rats to PM2.5 exposure. The result of general toxicology analysis revealed unconspicuous toxicity of PM2.5 under environmental dose, but winter PM2.5 at high dose caused severe histopathological damage to lung. Metabolomic analysis highlighted significant metabolic disorder induced by PM2.5 even at environmentally relevant doses. Lipid metabolism and GSH metabolism were primarily influenced by PM2.5 exposure due to the high levels of heavy metals. In addition, high levels of organic compounds such as PAHs, PCBs and PCDD/Fs in winter PM2.5 bring multiple overlaps on the toxic pathways, resulting in larger pulmonary toxicity and metabolic toxicity in rats than summer.

Do Connexin Mutants Cause Cataracts by Perturbing Glutathione Levels and Redox Metabolism in the Lens?

Cataracts of many different etiologies are associated with oxidation of lens components. The lens is protected by maintenance of a pool of reduced glutathione (GSH) and other antioxidants. Because gap junction channels made of the lens connexins, Cx46 and Cx50, are permeable to GSH, we tested whether mice expressing two different mutants, Cx46fs380 and Cx50D47A, cause cataracts by impairing lens glutathione metabolism and facilitating oxidative damage. Levels of GSH were not reduced in homogenates of whole mutant lenses. Oxidized glutathione (GSSG) and the GSSG/GSH ratio were increased in whole lenses of Cx50D47A, but not Cx46fs380 mice. The GSSG/GSH ratio was increased in the lens nucleus (but not cortex) of Cx46fs380 mice at 4.5 months of age, but it was not altered in younger animals. Carbonylated proteins were increased in Cx50D47A, but not Cx46fs380 lenses. Thus, both mouse lines have oxidizing lens environments, but oxidative modification is greater in Cx50D47A than in Cx46fs380 mice. The results suggest that GSH permeation through lens connexin channels is not a critical early event in cataract formation in these mice. Moreover, because oxidative damage was only detected in animals with significant cataracts, it cannot be an early event in their cataractogenesis.

Transporters and ascorbate-glutathione metabolism for differential cadmium accumulation and tolerance in two contrasting willow genotypes.

Salix matsudana Koidz is a low cadmium (Cd)-accumulating willow, whereas its cultivated variety, Salix matsudana var. matsudana f. umbraculifera Rehd., is a high Cd-accumulating and tolerant willow (HCW). The physiological and molecular mechanisms underlying differential Cd accumulation and tolerance in the two Salix species are poorly understood. Here, we confirmed that the differential Cd translocation capacity from roots to the shoots leads to the differential Cd accumulation in their aboveground parts between these two willow genotypes. Cadmium accumulation happens preferentially in the transport pathway, and Cd is mainly located in the vacuolar, cell wall and intercellular space in HCW bark by cadmium location analysis at tissue and subcellular levels. Comparative transcriptome analysis revealed that higher expressions of several metal transporter genes (ATP-binding cassette transporters, K+ transporters/channels, yellow stripe-like proteins, zinc-regulated transporter/iron-regulated transporter-like proteins, etc.) are involved in root uptake and translocation capacity in HCW; meanwhile, ascorbate-glutathione metabolic pathways play essential roles in Cd detoxification and higher tolerance of the Cd-accumulator HCW. These results lay the foundation for further understanding the molecular mechanisms of Cd accumulation in woody plants and provide new insights into molecular-assisted-screening woody plant varieties for phytoremediation.

Causal effects of serum metabolites on amyotrophic lateral sclerosis: A Mendelian randomization study.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder that is affected by both genetic and environmental factors. Nowadays, OMIC technologies, such as genomics and metabolomics, are providing a systematic readout of genetic structures and physiological states for understanding human diseases. However, the comprehensive analysis of cross-omics is often lacking. Here, we conducted a Mendelian randomization analysis to provide a comprehensive analysis of metabolomics and genomics to estimate the causal relationships between non-targeted human serum metabolites and the development of ALS. Using genetic variants as predictors, our study detected 18 metabolites that might have causal effects on the development of ALS, including a group of gamma-glutamyl amino acids. Our findings suggested that glutathione metabolism dysfunction might be involved in the pathogenesis of ALS. Furthermore, our study provides a novel method to understand the pathogenesis of human diseases and develop therapeutic strategies for diseases by combining metabolomics with genomics.

Glutathione Metabolism in Renal Cell Carcinoma Progression and Implications for Therapies.

A significantly increased level of the reactive oxygen species (ROS) scavenger glutathione (GSH) has been identified as a hallmark of renal cell carcinoma (RCC). The proposed mechanism for increased GSH levels is to counteract damaging ROS to sustain the viability and growth of the malignancy. Here, we review the current knowledge about the three main RCC subtypes, namely clear cell RCC (ccRCC), papillary RCC (pRCC), and chromophobe RCC (chRCC), at the genetic, transcript, protein, and metabolite level and highlight their mutual influence on GSH metabolism. A further discussion addresses the question of how the manipulation of GSH levels can be exploited as a potential treatment strategy for RCC.

Respiration disruption and detoxification at the protein expression levels in the Pacific oyster (Crassostrea gigas) under zinc exposure.

The Pacific oyster (Crassostrea gigas) can accumulate high levels of zinc (Zn) without obvious toxicity, but the related molecular mechanisms are largely unknown. In the present study, C. gigas were exposed to excess Zn for 9days and the differentially expressed proteins (DEPs) were examined using the isobaric tags for relative and absolute quantitation (iTRAQ) method. In total, 2667 proteins containing at least two peptides and detected in both replicates were used for proteomic analysis. Among these DEPs, 332 were up-regulated and 282 were down-regulated. KEGG enrichment analysis of DEPs revealed that Zn exposure mainly distrubed 'tricarboxylic acid (TCA) cycle', 'electron transport chain (ETC)' and 'glutathione (GSH) metabolism' processes in oysters. Further key protein expressions enriched in these metabolism pathways were analyzed. In TCA cycle, Zn inhibited the Fe-containing protein expressions, which may lead to the accumulation of succinate and induce anaerobiosis. In ETC metabolism process, Zn inhibited ETC complex protein expressions, including complex I-IV, which may affect the electron transport process. Furthermore, Zn induced phytochelatin (PC) and glutathione peroxidase (GPX) expression in GSH catabolism. The proteins play important roles in Zn detoxification and ROS elimination process. The transcriptional expressions of genes encoding these proteins were observed using real-time PCR analysis, and there was good consistency between these two datasets. Overall, we provide direct evidence for Zn toxicity and detoxification mechanisms at protein level.

Sulfur decreases cadmium translocation and enhances cadmium tolerance by promoting sulfur assimilation and glutathione metabolism in Brassica chinensis L.

We investigated the ameliorative role of sulfur (S) in protecting plants against cadmium (Cd) toxicity by using two pakchoi (Brassica chinensis L.) cultivars with different Cd tolerance levels. The exposure of pakchoi seedlings to 100µM Cd inhibited plant growth, increased superoxide content, enhanced membrane lipid peroxidation, and induced Cd accumulation in the roots and shoots. Application of S to Cd-stressed plants alleviated Cd-induced oxidative stress by promoting the capacity of the ascorbate (AsA)-glutathione (GSH) cycle, enhanced S assimilation by increasing the activity of ATP sulfurylase (ATPS) and o-acetylserine(thiol)lyase (OASTL), and decreased Cd translocation from the roots to the shoots by enhancing phytochelatins (PCs) biosynthesis. Results suggested that S reversed Cd-induced growth inhibition and oxidative stress by restraining Cd translocation from the roots to the shoots and upregulating S assimilation and GSH metabolism, including the AsA-GSH cycle and PCs synthesis.