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Lactate dehydrogenases amplify reactive oxygen species in cancer cells in response to oxidative stimuli.

ABSTRACT: Previous studies demonstrated that superoxide could initiate and amplify LDH-catalyzed hydrogen peroxide production in aqueous phase, but its physiological relevance is unknown. Here we showed that LDHA and LDHB both exhibited hydrogen peroxide-producing activity, which was significantly enhanced by the superoxide generated from the isolated mitochondria from HeLa cells and patients' cholangiocarcinoma specimen. After LDHA or LDHB were knocked out, hydrogen peroxide produced by Hela or 4T1 cancer cells were significantly reduced. Re-expression of LDHA in LDHA-knockout HeLa cells partially restored hydrogen peroxide production. In HeLa and 4T1 cells, LDHA or LDHB knockout or LDH inhibitor FX11 significantly decreased ROS induction by modulators of the mitochondrial electron transfer chain (antimycin, oligomycin, rotenone), hypoxia, and pharmacological ROS inducers piperlogumine (PL) and phenethyl isothiocyanate (PEITC). Moreover, the tumors formed by LDHA or LDHB knockout HeLa or 4T1 cells exhibited a significantly less oxidative state than those formed by control cells. Collectively, we provide a mechanistic understanding of a link between LDH and cellular hydrogen peroxide production or oxidative stress in cancer cells in vitro and in vivo.

SUBMITTER: Wu H

PROVIDER: S-EPMC8236487 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Cells have distinct mechanisms to maintain protection against different reactive oxygen species: oxidative-stress-response genes.

Project description:The complete set of viable deletion strains in Saccharomyces cerevisiae was screened for sensitivity of mutants to five oxidants to identify cell functions involved in resistance to oxidative stress. This screen identified a unique set of mainly constitutive functions providing the first line of defense against a particular oxidant; these functions are very dependent on the nature of the oxidant. Most of these functions are distinct from those involved in repair and recovery from damage, which are generally induced in response to stress, because there was little correlation between mutant sensitivity and the reported transcriptional response to oxidants of the relevant gene. The screen identified 456 mutants sensitive to at least one of five different types of oxidant, and these were ranked in order of sensitivity. Many genes identified were not previously known to have a role in resistance to reactive oxygen species. These encode functions including protein sorting, ergosterol metabolism, autophagy, and vacuolar acidification. Only two mutants were sensitive to all oxidants examined, only 12 were sensitive to at least four, and different oxidants had very different spectra of deletants that were sensitive. These findings highlight the specificity of cellular responses to different oxidants: No single oxidant is representative of general oxidative stress. Mitochondrial respiratory functions were overrepresented in mutants sensitive to H(2)O(2), and vacuolar protein-sorting mutants were enriched in mutants sensitive to diamide. Core functions required for a broad range of oxidative-stress resistance include transcription, protein trafficking, and vacuolar function.

| S-EPMC404085 | biostudies-literature

Lactate regulates rat male germ cell function through reactive oxygen species.

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| S-EPMC3909278 | biostudies-literature

Human Leukemic Cells performing Oxidative Phosphorylation (OXPHOS) Generate an Antioxidant Response Independently of Reactive Oxygen species (ROS) Production.

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| S-EPMC4739420 | biostudies-literature

Reactive Oxygen Species Imaging in U937 Cells.

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| S-EPMC7593787 | biostudies-literature

Reactive Oxygen Species Differentially Modulate the Metabolic and Transcriptomic Response of Endothelial Cells.

Project description:Reactive oxygen species (ROS) are important mediators of both physiological and pathophysiological signal transduction in the cardiovascular system. The effects of ROS on cellular processes depend on the concentration, localization, and duration of exposure. Cellular stress response mechanisms have evolved to mitigate the negative effects of acute oxidative stress. In this study, we investigate the short-term and long-term metabolic and transcriptomic response of human umbilical vein endothelial cells (HUVEC) to different types and concentrations of ROS. To generate intracellular H2O2, we utilized a lentiviral chemogenetic approach for overexpression of human D-amino acid oxidase (DAO). DAO converts D-amino acids into their corresponding imino acids and H2O2. HUVEC stably overexpressing DAO (DAO-HUVEC) were exposed to D-alanine (3 mM), exogenous H2O2 (10 µM or 300 µM), or menadione (5 µM) for various timepoints and subjected to global untargeted metabolomics (LC-MS/MS) and RNAseq by MACE (Massive analysis of cDNA ends). A total of 300 µM H2O2 led to pronounced changes on both the metabolic and transcriptomic level. In particular, metabolites linked to redox homeostasis, energy-generating pathways, and nucleotide metabolism were significantly altered. Furthermore, 300 µM H2O2 affected genes related to the p53 pathway and cell cycle. In comparison, the effects of menadione and DAO-derived H2O2 mainly occurred at gene expression level. Collectively, all types of ROS led to subtle changes in the expression of ribosomal genes. Our results show that different types and concentration of ROS lead to a different metabolic and transcriptomic response in endothelial cells.

| S-EPMC8869421 | biostudies-literature

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Project description:Antioxidants play an important role in regulating overabundant reactive oxygen species (ROS) in wound healing to reduce oxidative stress and inflammation. In this work, we demonstrate for the first time that functionalization of methacrylamide chitosan (MAC) with aliphatic pentadecafluoro chains, to synthesize pentadecafluoro-octanoyl methacrylamide chitosan (MACF), enhances the antioxidant capacity of the MAC base hydrogel material, while being able to deliver oxygen for future enhanced wound healing applications. As such, MACF was shown to sequester more nitric oxide (p < 0.01) and hydroxyl (p < 0.0001) radicals as compared to the negative control even when delivering additional oxygen. MACF's beneficial antioxidant capacity was further confirmed in in vitro cell culture experiments using human dermal fibroblasts stressed with 2,2'-azobis(2-methylpropionamidine) dihydrochloride (AAPH). © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2368-2374, 2017.

| S-EPMC5533114 | biostudies-literature

Structurally Linked Dynamics in Lactate Dehydrogenases of Evolutionarily Distinct Species.

Project description:We present new findings about how primary and secondary structure affects the role of fast protein motions in the reaction coordinates of enzymatic reactions. Using transition path sampling and committor distribution analysis, we examined the difference in the role of these fast protein motions in the reaction coordinate of lactate dehydrogenases (LDHs) of Apicomplexa organisms Plasmodium falciparum and Cryptosporidium parvum. Having evolved separately from a common malate dehydrogenase ancestor, the two enzymes exhibit several important structural differences, notably a five-amino acid insertion in the active site loop of P. falciparum LDH. We find that these active site differences between the two organisms' LDHs likely cause a decrease in the contribution of the previously determined LDH rate-promoting vibration to the reaction coordinate of P. falciparum LDH compared to that of C. parvum LDH, specifically in the coupling of the rate-promoting vibration and the hydride transfer. This effect, while subtle, directly shows how changes in structure near the active site of LDH alter catalytically important motions. Insights provided by studying these alterations would prove to be useful in identifying LDH inhibitors that specifically target the isozymes of these parasitic organisms.

| S-EPMC5507060 | biostudies-literature

ATP activates a reactive oxygen species-dependent oxidative stress response and secretion of proinflammatory cytokines in macrophages.

Project description:Secretion of the proinflammatory cytokines, interleukin (IL)-1beta and IL-18, usually requires two signals. The first, due to microbial products such as lipopolysaccharide, initiates transcription of the cytokine genes and accumulation of the precursor proteins. Cleavage and secretion of the cytokines is mediated by caspase-1, in association with an inflammasome containing Nalp3, which can be activated by binding of extracellular ATP to purinergic receptors. We show that treatment of macrophages with ATP results in production of reactive oxygen species (ROS), which stimulate the phosphatidylinositol 3-kinase (PI3K) pathway and subsequent Akt and ERK1/2 activation. ROS exerts its effect through glutathionylation of PTEN (phosphatase and tensin homologue deleted from chromosome 10), whose inactivation would shift the equilibrium in favor of PI3K. ATP-dependent ROS production and PI3K activation also stimulate transcription of genes required for an oxidative stress response. In parallel, ATP-mediated ROS-dependent PI3K is required for activation of caspase-1 and secretion of IL-1beta and IL-18. Thus, an increase in ROS levels in ATP-treated macrophages results in activation of a single pathway that promotes both adaptation to subsequent exposure to oxidants or inflammation, and processing and secretion of proinflammatory cytokines.

| S-EPMC2693903 | biostudies-literature

Increased Reactive Oxygen Species Formation and Oxidative Stress in Rheumatoid Arthritis.

Project description:BackgroundRheumatoid arthritis (RA) is an autoimmune inflammatory disorder. Highly reactive oxygen free radicals are believed to be involved in the pathogenesis of the disease. In this study, RA patients were sub-grouped depending upon the presence or absence of rheumatoid factor, disease activity score and disease duration. RA Patients (120) and healthy controls (53) were evaluated for the oxidant-antioxidant status by monitoring ROS production, biomarkers of lipid peroxidation, protein oxidation and DNA damage. The level of various enzymatic and non-enzymatic antioxidants was also monitored. Correlation analysis was also performed for analysing the association between ROS and various other parameters.MethodsIntracellular ROS formation, lipid peroxidation (MDA level), protein oxidation (carbonyl level and thiol level) and DNA damage were detected in the blood of RA patients. Antioxidant status was evaluated by FRAP assay, DPPH reduction assay and enzymatic (SOD, catalase, GST, GR) and non-enzymatic (vitamin C and GSH) antioxidants.ResultsRA patients showed a higher ROS production, increased lipid peroxidation, protein oxidation and DNA damage. A significant decline in the ferric reducing ability, DPPH radical quenching ability and the levels of antioxidants has also been observed. Significant correlation has been found between ROS and various other parameters studied.ConclusionRA patients showed a marked increase in ROS formation, lipid peroxidation, protein oxidation, DNA damage and decrease in the activity of antioxidant defence system leading to oxidative stress which may contribute to tissue damage and hence to the chronicity of the disease.

| S-EPMC4820274 | biostudies-literature

Multivalent Ligand-Nanoparticle Conjugates Amplify Reactive Oxygen Species Second Messenger Generation and Enhance Epidermal Growth Factor Receptor Phosphorylation.

Project description:The epidermal growth factor (EGF) receptor (EGFR) is heterogeneously distributed on the cellular surface and enriched in clusters with diameters of tens of nanometers. Multivalent presentation of EGF ligand on nanoparticles (NPs) provides an approach for controlling and amplifying the local activation of EGFR in these clusters. Reactive oxygen species (ROS) have been indicated to play a role in the regulation of EGFR activation as second messengers, but the effect of nanoconjugation on EGF-mediated ROS formation and ROS-induced EGFR activation is not well established. The goal of this manuscript is to characterize the multivalent enhancement of EGF-induced ROS formation and to test its effect on EGFR phosphorylation in breast cancer cell models using gold (Au) NPs with a diameter of 81 ± 1 nm functionalized with two different EGF ligand densities (12 ± 7 EGF/NP (NP-EGF12) and 87 ± 6 EGF/NP (NP-EGF87)). In the EGFR overexpressing cell lines MDA-MB-231 and MDA-MB-468, NP-EGF87 achieved a measurable multivalent enhancement of ROS that peaked at concentrations c ROSmax ≤ 25 pM and that were EGFR and nicotinamide adenine dinucleotide phosphate oxidase (NOX) dependent. NP-EGF12 failed to generate comparable ROS levels as NP-EGF87 in the investigated NP input concentration range (0-100 pM). In cells with nearly identical numbers of bound NP-EGF87 and NP-EGF12, the ROS levels for NP-EGF87 were systematically higher, indicating that the multivalent enhancement is exclusively related not only to avidity but also to a stronger stimulation per NP. Importantly, the increase in EGF-induced ROS formation associated with EGF nanoconjugation at c ROSmax resulted in a measurable gain in EGFR phosphorylation, confirming that ROS generation contributes to the multivalent enhancement of EGFR activation in response to NP-EGF87.

| S-EPMC9815836 | biostudies-literature

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