scholarly journals Vitamin C Is a Kinase Inhibitor: Dehydroascorbic Acid Inhibits IκBα Kinase β

2004 ◽  
Vol 24 (15) ◽  
pp. 6645-6652 ◽  
Author(s):  
Juan M. Cárcamo ◽  
Alicia Pedraza ◽  
Oriana Bórquez-Ojeda ◽  
Bing Zhang ◽  
Roberto Sanchez ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Vitamin C ◽  
Enzymatic Activity ◽  
Redox State ◽  
Kinase Inhibitor ◽  
Dehydroascorbic Acid ◽  
Molecular Action ◽  
Cellular Signal ◽  
Direct Linkage

ABSTRACT Reactive oxygen species (ROS) are key intermediates in cellular signal transduction pathways whose function may be counterbalanced by antioxidants. Acting as an antioxidant, ascorbic acid (AA) donates two electrons and becomes oxidized to dehydroascorbic acid (DHA). We discovered that DHA directly inhibits IκBα kinase β (IKKβ) and IKKα enzymatic activity in vitro, whereas AA did not have this effect. When cells were loaded with AA and induced to generate DHA by oxidative stress in cells expressing a constitutive active IKKβ, NF-κB activation was inhibited. Our results identify a dual molecular action of vitamin C in signal transduction and provide a direct linkage between the redox state of vitamin C and NF-κB signaling events. AA quenches ROS intermediates involved in the activation of NF-κB and is oxidized to DHA, which directly inhibits IKKβ and IKKα enzymatic activity. These findings define a function for vitamin C in signal transduction other than as an antioxidant and mechanistically illuminate how vitamin C down-modulates NF-κB signaling.

Vitamin C Recycling Regulates Neurite Growth in Neurospheres Differentiated in Vitro

2020 ◽  
Author(s):  
Francisca Espinoza ◽  
Rocio Magdalena ◽  
Natalia Saldivia ◽  
Nery Jara ◽  
Fernando Martínez ◽  
...  
Keyword(s):  
Vitamin C ◽  
Bystander Effect ◽  
Dehydroascorbic Acid ◽  
Neurite Growth ◽  
Protein Carbonylation ◽  
Reduced Form ◽  
Ros Generation ◽  
Lysine Production ◽  
Neuritic Growth

Abstract Background: The reduced form of vitamin C, ascorbic acid (AA), has been related to antioxidant defense as well as gene expression and cell differentiation in the cerebral cortex. In neurons, AA is mainly oxidized to dehydroascorbic acid (DHA); however, DHA cannot accumulate intracellularly because it induces metabolic changes and cell death. In this context, it has been proposed that vitamin C recycling via neuron-astrocyte coupling maintains AA levels and prevents DHA parenchymal accumulation. To date, the role of this mechanism during the outgrowth of neurites is unknown.Methods: To stimulate neuronal differentiation, adhered neurospheres treated with AA and retinoic acid (RA) were used. Neuritic growth was analyzed by confocal microscopy, and the effect of vitamin C recycling (bystander effect) in vitro was studied using different cells (astrocytes, HL60 and U87). Reactive oxygen species (ROS) generation was also analyzed by flow cytometry and protein carbonylation / carboximetil-lysine production.Results: AA stimulates neuritic growth more efficiently than RA. However, AA is oxidized to DHA in long incubation periods, generating a loss in the formation of neurites. Surprisingly, neurite growth is maintained over time following co-incubation of neurospheres with cells (HL60, U87, or astrocytes) that efficiently capture DHA (Bystander effect). In this sense, astrocytes have high capacity to recycle DHA and stimulate the maintenance of neurites. Finally, our data indicate that DHA induces ROS generation, a condition that results in protein carbonylation and carboximetil-lysine production. Conclusions: We have demonstrated that vitamin C recycling in vitro regulates the morphology of immature neurons during the differentiation and maturation processes.

scholarly journals Selective inhibition of Ph-positive ALL cell growth through kinase-dependent and -independent effects by CDK6-specific PROTACs

Blood ◽  
2020 ◽  
Vol 135 (18) ◽  
pp. 1560-1573 ◽  
Author(s):  
Marco De Dominici ◽  
Patrizia Porazzi ◽  
Youcai Xiao ◽  
Allen Chao ◽  
Hsin-Yao Tang ◽  
...  
Keyword(s):  
Cell Growth ◽  
Enzymatic Activity ◽  
Kinase Inhibitor ◽  
De Novo ◽  
Lymphoblastic Leukemia ◽  
Regulatory Gene ◽  
Selective Inhibition ◽  
Hematologic Malignancies ◽  
Hematopoietic Progenitors

Abstract Expression of the cell cycle regulatory gene CDK6 is required for Philadelphia-positive (Ph+) acute lymphoblastic leukemia (ALL) cell growth, whereas expression of the closely related CDK4 protein is dispensable. Moreover, CDK6 silencing is more effective than treatment with the dual CDK4/6 inhibitor palbociclib in suppressing Ph+ ALL in mice, suggesting that the growth-promoting effects of CDK6 are, in part, kinase-independent in Ph+ ALL. Accordingly, we developed CDK4/6–targeted proteolysis-targeting chimeras (PROTACs) that inhibit CDK6 enzymatic activity in vitro, promote the rapid and preferential degradation of CDK6 over CDK4 in Ph+ ALL cells, and markedly suppress S-phase cells concomitant with inhibition of CDK6-regulated phospho-RB and FOXM1 expression. No such effects were observed in CD34+ normal hematopoietic progenitors, although CDK6 was efficiently degraded. Treatment with the CDK6-degrading PROTAC YX-2-107 markedly suppressed leukemia burden in mice injected with de novo or tyrosine kinase inhibitor–resistant primary Ph+ ALL cells, and this effect was comparable or superior to that of the CDK4/6 enzymatic inhibitor palbociclib. These studies provide “proof of principle” that targeting CDK6 with PROTACs that inhibit its enzymatic activity and promote its degradation represents an effective strategy to exploit the “CDK6 dependence” of Ph+ ALL and, perhaps, of other hematologic malignancies. Moreover, they suggest that treatment of Ph+ ALL with CDK6-selective PROTACs would spare a high proportion of normal hematopoietic progenitors, preventing the neutropenia induced by treatment with dual CDK4/6 inhibitors.

scholarly journals Bioaccessibility of Antioxidants in Blackcurrant Juice After Treatment Using Supercritical Carbon Dioxide

2021 ◽  
Author(s):  
Urszula Trych ◽  
Magdalena Buniowska ◽  
Sylwia Skąpska ◽  
Ireneusz Kapusta ◽  
Krystian Marszałek
Keyword(s):  
Carbon Dioxide ◽  
Supercritical Carbon Dioxide ◽  
Vitamin C ◽  
Model Experiment ◽  
Dehydroascorbic Acid ◽  
Total Anthocyanins ◽  
The Stability ◽  
Supercritical Carbon ◽  
Thermally Treated

Blackcurrant juice (Ribes nigrum L.) was subjected to supercritical carbon dioxide (SCCD) at 10, 30 and 60 MPa for 10 min at 45°C as well as thermally treated at 45°C and 85°C for 10 min to determine the stability, antioxidant capacity (AC) and bioaccessibility (BAc) of vitamin C, total anthocyanins and their individual monomers. An in vitro gastrointestinal digestion model completed with dialysis was used to assess BAc. The use of SCCD at each of the pressures applied improved the stability of vitamin C, total anthocyanins, and AC before in vitro digestion. As a result of digestion, L-ascorbic acid was oxidized to L-dehydroascorbic acid, and finally, the total content of vitamin C, anthocyanins, and AC decreased. SCCD did not significantly improve the BAc of vitamin C and total anthocyanins. The highest BAc of vitamin C was noted in fresh juice (FJ) (40%) and after mild heat treatment at 45°C (T45) (46%). The highest BAc of total anthocyanins was also noted in the FJ (4.4%). The positive effect of the application of SCCD on the BAc of the delphinidin-3-O-glycosides was observed compared to T45 and thermal pasteurization at 85°C (T85). Moreover, cyanidins were generally more bioaccessible than delphinidins in all samples. AC after digestion was higher in SCCD samples compared to thermally treated measured using ABTS+• and DPPH• assays, whereas in dialysate similar trends were observed only for AC measured using the ABTS+• assay. This phenomenon was justified by the formation of individual metabolites detected by UPLC-PDA-MS / MS in the model experiment with delphinidin-3-O-rutinoside. The protocatechuic acid which is well known as a strong antioxidant was detected in the model experiment after digestion. Further research is needed to better understand the metabolic pathway of anthocyanins and the possible uses of SCCD to improve the health properties of fruit products.

scholarly journals Effects of Ozone Oxidative Preconditioning on TNF-αRelease and Antioxidant-Prooxidant Intracellular Balance in Mice During Endotoxic Shock

2005 ◽  
Vol 2005 (1) ◽  
pp. 16-22 ◽  
Author(s):  
Zullyt B. Zamora ◽  
Aluet Borrego ◽  
Orlay Y. López ◽  
René Delgado ◽  
Ricardo González ◽  
...  
Keyword(s):  
Glutathione Peroxidase ◽  
Enzymatic Activity ◽  
Redox State ◽  
Endotoxic Shock ◽  
Gaseous Mixture ◽  
Hepatic Tissue ◽  
Mouse Serum ◽  
Antioxidant Systems ◽  
Dependent Manner

Ozone oxidative preconditioning is a prophylactic approach, which favors the antioxidant-prooxidant balance for preservation of cell redox state by the increase of antioxidant endogenous systems in both in vivo and in vitro experimental models. Our aim is to analyze the effect of ozone oxidative preconditioning on serum TNF-αlevels and as a modulator of oxidative stress on hepatic tissue in entodoxic shock model (mice treated with lipopolysaccharide (LPS)). Ozone/oxygen gaseous mixture which was administered intraperitoneally (0.2,0.4, and1.2mg/kg) once daily for five days before LPS (0.1mg/kg, intraperitoneal). TNF-αwas measured by cytotoxicity on L-929 cells. Biochemical parameters such as thiobarbituric acid reactive substances (TBARS), enzymatic activity of catalase, glutathione peroxidase, and glutathione-S transferase were measured in hepatic tissue. One hour after LPS injection there was a significant increase in TNF-αlevels in mouse serum. Ozone/oxygen gaseous mixture reduced serum TNF-αlevels in a dose-dependent manner. Statistically significant decreases in TNF-αlevels after LPS injection were observed in mice pretreated with ozone intraperitoneal applications at0.2(78%),0.4(98%), and1.2(99%). Also a significant increase in TBARS content was observed in the hepatic tissue of LPS-treated mice, whereas enzymatic activity of glutathion-S transferase and glutathione peroxidase was decreased. However in ozone-treated animals a significant decrease in TBARS content was appreciated as well as an increase in the activity of antioxidant enzymes. These results indicate that ozone oxidative preconditioning exerts inhibitory effects on TNF-αproduction and on the other hand it exerts influence on the antioxidant-prooxidant balance for preservation of cell redox state by the increase of endogenous antioxidant systems.

scholarly journals Alternative pathways of dehydroascorbic acid degradation in vitro and in plant cell cultures: novel insights into vitamin C catabolism

2011 ◽  
Vol 440 (3) ◽  
pp. 375-385 ◽  
Author(s):  
Harriet T. Parsons ◽  
Tayyaba Yasmin ◽  
Stephen C. Fry
Keyword(s):  
Reactive Oxygen Species ◽  
Vitamin C ◽  
Dehydroascorbic Acid ◽  
Oxygen Species ◽  
Reactive Intermediate ◽  
Species Status ◽  
Alternative Pathways ◽  
Irreversible Oxidation

L-Ascorbate catabolism involves reversible oxidation to DHA (dehydroascorbic acid), then irreversible oxidation or hydrolysis. The precursor–product relationships and the identity of several major DHA breakdown products remained unclear. In the presence of added H2O2, DHA underwent little hydrolysis to DKG (2,3-dioxo-L-gulonate). Instead, it yielded OxT (oxalyl L-threonate), cOxT (cyclic oxalyl L-threonate) and free oxalate (~6:1:1), essentially simultaneously, suggesting that all three product classes independently arose from one reactive intermediate, proposed to be cyclic-2,3-O-oxalyl-L-threonolactone. Only with plant apoplastic esterases present were the esters significant precursors of free oxalate. Without added H2O2, DHA was slowly hydrolysed to DKG. Downstream of DKG was a singly ionized dicarboxy compound (suggested to be 2-carboxy-L-xylonolactone plus 2-carboxy-L-lyxonolactone), which reversibly de-lactonized to a dianionic carboxypentonate. Formation of these lactones and acid was minimized by the presence of residual unreacted ascorbate. In vivo, the putative 2-carboxy-L-pentonolactones were relatively stable. We propose that DHA is a branch-point in ascorbate catabolism, being either oxidized to oxalate and its esters or hydrolysed to DKG and downstream carboxypentonates. The oxidation/hydrolysis ratio is governed by reactive oxygen species status. In vivo, oxalyl esters are enzymatically hydrolysed, but the carboxypentonates are stable. The biological roles of these ascorbate metabolites invite future exploration.

scholarly journals LipA, a Tyrosine and Lipid Phosphatase Involved in the Virulence of Listeria monocytogenes

2011 ◽  
Vol 79 (6) ◽  
pp. 2489-2498 ◽  
Author(s):  
Renate Kastner ◽  
Olivier Dussurget ◽  
Cristel Archambaud ◽  
Elisabeth Kernbauer ◽  
Didier Soulat ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Listeria Monocytogenes ◽  
Enzymatic Activity ◽  
Phosphatase Activity ◽  
Content Type ◽  
Conserved Motif ◽  
Cell Functions ◽  
Lipid Phosphatase

ABSTRACTIntracellular bacterial pathogens manipulate host cell functions by producing enzymes that stimulate or antagonize signal transduction. TheListeria monocytogenesgenome contains a gene,lmo1800, encoding a protein with a conserved motif of conventional tyrosine phosphatases. Here, we report that thelmo1800-encoded protein LipA is secreted byListeriaand displays tyrosine as well as lipid phosphatase activityin vitro. Bacteria lacking LipA are severely attenuated in virulencein vivo, thus revealing a so-far-undescribed enzymatic activity involved inListeriainfection.

scholarly journals The serine/threonine kinase MINK1 directly regulates the function of promigratory proteins

2021 ◽  
Author(s):  
Avais Daulat ◽  
Monica Silveira Wagner ◽  
Stephane Audebert ◽  
Malgorzata Kowalczewska ◽  
Jeremy Ariey-Bonnet ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Protein Complex ◽  
Planar Cell Polarity ◽  
Kinase Inhibitor ◽  
Cell Cortex ◽  
Mrna Levels ◽  
Serine Threonine Kinase ◽  
Threonine Kinase

Upregulation of the developmental Wnt/planar cell polarity pathway is observed in many cancers and is associated with cancer development at early and late stages. We recently showed that PRICKLE1 and VANGL2, two core Wnt/PCP components, are overexpressed in triple negative breast cancer and associated with poor prognosis. PRICKLE1 is a cytoplasmic protein phosphorylated by the poorly described serine/threonine kinase MINK1 which triggers its localization at the plasma membrane, a key step for its function. Knockdown experiments have demonstrated that MINK1 and PRICKLE1 contribute to TNBC cell motility and spreading in vitro and in vivo. However, the identity of MINK1 substrates and the role of MINK1 enzymatic activity in this process have not yet been addressed issues. We carried out a phosphoproteomic strategy and identified novel MINK1 substrates including LL5β. LL5β is a membrane scaffold molecule that anchors microtubules at the cell cortex through its association with the plus-end MT proteins CLASPs to trigger focal adhesion disassembly. LL5β is a prominent member of the MINK1-PRICKLE1 protein complex and is directly phosphorylated by MINK1 that promotes its interaction with CLASP. Using a kinase inhibitor, we demonstrate that the enzymatic activity of MINK1 is involved in the protein complex assembly and localization, and cell migration. Analysis of gene expression data show that the concomitant up-regulation of PRICKLE1 and LL5β mRNA levels encoding MINK1 substrates is associated with a poor metastasis-free survival for TNBC patients. Altogether, our results suggest that MINK1 may represent a potential target in TNBC.

scholarly journals Sirt2-SUMOylation is essential for its tumor-suppressor function in neuroblastoma

2020 ◽  
Author(s):  
Wenmei Lu ◽  
Qian Wang ◽  
Ci Xu ◽  
Haihua Yuan ◽  
Qiang Fan ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Tumor Suppressor ◽  
Survival Curve ◽  
Neuroblastoma Cell ◽  
Specific Inhibitor ◽  
Migration And Invasion ◽  
Cellular Signal Transduction ◽  
Cellular Signal

Abstract Background SUMOylation is an important post-translational modification and participates in a variety of cellular physiological and pathological processes in eukaryotic cells. Sirt2, as a NAD+-dependent deacetylase, usually exerts tumor-suppressor function. However, its SUMOylation roles in cancer cells have not been illustrated. Methods Ni2+-NTA, GST-pull down and immunoprecipitation in vitro and in vivo were used for the Sirt2-SUMOylation or P38-Acetylation assay. Immunofluorescence and Nuclear/Cytosol Fractionation Kit were performed to identify the localization of Sirt2 with or without SUMOylation. The proliferation, soft-agar colony formation, migration and invasion were performed to detect the phenotypes of neuroblastoma cells in vitro, and the xenograft tumor model was conducted to verify Sirt2-SUMOylation role in tumorigenesis in vivo. R2 online database were used for the clinical analysis, including expression, survival curve and pathology stage. Results SUMOylation can occur in Sirt2 protein at both lysine 183 and lysine 340 sites. SUMOylation of Sirt2 does not affect its localization or stability, but involves in the P38-mTORC2-AKT cellular signal transduction via the direct deacetylation on P38. SUMOylation-deficient Sirt2 losses the capability of suppressing tumor processes and neuroblastoma cell shows a resistance to Sirt2-specific inhibitor AK-7. Conclusion We revealed an important function of SUMOylation on Sirt2, which is closely associated with the cellular signal transduction and is essential for suppressing the tumorigenesis in neuroblastoma.

scholarly journals Vitamin C protects HL60 and U266 cells from arsenic toxicity

Blood ◽  
2005 ◽  
Vol 105 (10) ◽  
pp. 4004-4012 ◽  
Author(s):  
Nicos Karasavvas ◽  
Juan M. Cárcamo ◽  
George Stratis ◽  
David W. Golde
Keyword(s):  
Vitamin C ◽  
Culture Media ◽  
Dehydroascorbic Acid ◽  
Transition Metal Ions ◽  
Glutathione Depletion ◽  
Apparent Paradox ◽  
Free Transition ◽  
Rpmi 8226 ◽  
Dose Dependent

AbstractAlthough there is no compelling evidence that vitamin C has antitumor activity in humans, clinical trials are testing the hypothesis that ascorbic acid (AA) will enhance the efficacy of arsenic trioxide (As2O3) in myeloma. In vitro, AA cytotoxicity depends on its interaction with free transition metal ions in culture media leading to the generation of H2O2 and other reactive oxygen species (ROSs). Therefore, to circumvent the extracellular in vitro pro-oxidant effects of AA, we loaded HL60, U266, and RPMI-8226 cells with vitamin C by incubation with dehydroascorbic acid (DHA). Loading cells in this manner resulted in prominent, dose-dependent protection of As2O3-treated cells as measured by viability, colony formation, and apoptosis assays. Glutathione depletion enhanced cell sensitivity to the cytotoxic effects of As2O3 and vitamin C loading provided protection. AA was found to generate cytotoxic concentrations of H2O2 in culture medium without cells and copper/iron chelators inhibited this reaction. However, AA did not generate H2O2 in simple buffer or human plasma. Direct incubation with AA resulted in increased intracellular ROSs, whereas DHA incubation decreased it. These results clarify an apparent paradox and indicate that vitamin C loading in HL60, U266, and RPMI-8226 cells ameliorates As2O3 cytotoxicity.

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