RESUMEN

Since the discovery of ferroptosis, it has been postulated that this type of cell death could be utilized in treatments for cancer. Unfortunately, several highly aggressive tumor models are resistant to the pharmacological induction of ferroptosis. However, with the use of combined therapies, it is possible to recover sensitivity to ferroptosis in certain cellular models. Here, we discovered that co-treatment with the metabolically stable ferroptosis inducer imidazole ketone erastin (IKE) and the oxidized form of vitamin C, dehydroascorbic acid (DHAA), is a powerful therapy that induces ferroptosis in tumor cells previously resistant to IKE-induced ferroptosis. We determined that DHAA and IKE + DHAA delocalize and deplete GPX4 in tumor cells, specifically inducing lipid droplet peroxidation, which leads to ferroptosis. Moreover, in vivo, IKE + DHAA has high efficacy with regard to the eradication of highly aggressive tumors such as glioblastomas. Thus, the use of IKE + DHAA could be an effective and safe therapy for the eradication of difficult-to-treat cancers.

Asunto(s)

Ferroptosis , Neoplasias , Humanos , Ácido Deshidroascórbico/farmacología , Gotas Lipídicas , Muerte Celular , Peroxidación de Lípido

RESUMEN

Black spot rot caused by Alternaria alternata is one of the major postharvest disease of apple fruit during logistic. This study evaluated in vitro inhibitory effect of 2-hydroxy-3-phenylpropanoic acid (PLA) at various concentrations on A. alternata and the possible mechanisms involved in its action. Results showed that different concentrations of PLA inhibited conidia germination and mycelial growth of A. alternata in vitro, and 1.0 g L-1 was the lowest effective concentration to suppress A. alternata growth. Moreover, PLA significantly reduced relative conductivity and increased malondialdehyde and soluble protein contents. PLA also increased H2O2 and dehydroascorbic acid contents, but reduced ascorbic acid content. Additionally, PLA treatment inhibited catalase, ascorbate peroxidase, monodehydroascorbate acid reductase, dehydroascorbic acid reductase and glutathione reductase activities, whereas promoted superoxide dismutase activity. All these findings suggest that the possible mechanisms involved in the inhibitory effect of PLA on A. alternata included damaging the cell membrane integrity to cause electrolyte leakage and destroying reactive oxygen species balance.

Asunto(s)

Ácido Deshidroascórbico , Peróxido de Hidrógeno , Especies Reactivas de Oxígeno/metabolismo , Peróxido de Hidrógeno/farmacología , Ácido Deshidroascórbico/farmacología , Membrana Celular , Alternaria , Poliésteres/farmacología

RESUMEN

Aims: Glioblastoma (GB) is one of the most aggressive brain tumors. These tumors modify their metabolism, increasing the expression of glucose transporters, GLUTs, which incorporate glucose and the oxidized form of vitamin C, dehydroascorbic acid (DHA). We hypothesized that GB cells preferentially take up DHA, which is intracellularly reduced and compartmentalized into the endoplasmic reticulum (ER), promoting collagen biosynthesis and an aggressive phenotype. Results: Our results showed that GB cells take up DHA using GLUT1, while GLUT3 and sodium-dependent vitamin C transporter 2 (SVCT2) are preferably intracellular. Using a baculoviral system and reticulum-enriched extracts, we determined that SVCT2 is mainly located in the ER and corresponds to a short isoform. Ascorbic acid (AA) was compartmentalized, stimulating collagen IV secretion and increasing in vitro and in situ cell migration. Finally, orthotopic xenografts induced in immunocompetent guinea pigs showed that vitamin C deficiency retained collagen, reduced blood vessel invasion, and affected glomeruloid vasculature formation, all pathological conditions associated with malignancy. Innovation and Conclusion: We propose a functional role for vitamin C in GB development and progression. Vitamin C is incorporated into the ER of GB cells, where it favors the synthesis of collagen, thus impacting tumor development. Collagen secreted by tumor cells favors the formation of the glomeruloid vasculature and enhances perivascular invasion. Antioxid. Redox Signal. 37, 538-559.

Asunto(s)

Ácido Ascórbico , Glioblastoma , Animales , Ácido Ascórbico/metabolismo , Ácido Ascórbico/farmacología , Colágeno/metabolismo , Ácido Deshidroascórbico/metabolismo , Ácido Deshidroascórbico/farmacología , Glucosa/metabolismo , Cobayas , Humanos , Transportadores de Sodio Acoplados a la Vitamina C/metabolismo , Vitaminas

RESUMEN

Efforts to develop effective drugs targeting PI3K and KRAS signaling pathways in PIK3CA/KRAS-mutant colorectal cancer stem cells (CRCSCs) remain challenging. Finding safe compounds that can easily enter CRCSCs with the ability to target metastasis-driver gene CXCR4 and pluripotency network genes as key upstream and downstream effectors of both PI3K and KRAS signaling pathways may provide promising results. PIK3CA/KRAS-mutant CRCSCs display high expression of glucose transporters (GLUTs) on their cell membrane and a glycolytic phenotype providing an opportunity to deliver antiglycolytic compounds into these cells via the GLUTs. CRC patients with low levels of vitamin C in their plasma show a shorter survival suggesting the ability of this vitamin at the physiologic levels for caspase-3 activation and apoptosis in CRCSCs. Vitamin C in an oxidized form (L-dehydroascorbic acid; L-DHA) with antiglycolytic activity can be taken up into CRC cells via the GLUTs. This may provide selective toxicity on CRCSCs and affect CXCR4 and stemness markers genes expression in these cells. To this end, we treated PIK3CA/KRAS-mutant LS174T cells with high glycolytic activity as an attractive model for CRCSCs with L-DHA equal to the pharmacological levels of vitamin C in human plasma, after which cell numbers, metabolic activity, proliferation-rate, CXCR4 and pluripotency network genes expression, caspase-3 activity with apoptosis were evaluated. 48 h post-treatment with 100- to 1000 µM L-DHA, cell numbers were decreased and measured to be 70-47% control. L-DHA with selective toxicity on LS174T cells diminished metabolic activity and cell proliferation-rate to 1.4-0.8 (Control OD = 1.5) and 92-54.5% respectively with no toxicity on PBMCs. L-DHA decreased CXCR4, Bmi-1, Sox-2 and Oct-4 expression to 45%, 85%, 45% and 48% control respectively followed by caspase-3 reactivation by 2.5 to 4.9-fold increases and induction of apoptosis ranging from 0.5% to 58.3% for 100- to 1000 µM L-DHA. According to our data, CRC stem-like cells were highly sensitive to L-DHA in in-vitro. L-DHA selectively targeted LS174T cells and successfully reactivated caspase-3 and apoptosis in these cells. CXCR4, stemness marker genes and metabolic activity appear to be promising targets of L-DHA. Our results may provide a new therapeutic approach to target selectively GLUT-overexpressing PIK3CA/KRAS-mutant CRCSCs using L-DHA with no toxicity on normal cells.

Asunto(s)

Neoplasias Colorrectales , Fosfatidilinositol 3-Quinasas , Línea Celular Tumoral , Fosfatidilinositol 3-Quinasa Clase I/metabolismo , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Ácido Deshidroascórbico/farmacología , Ácido Deshidroascórbico/uso terapéutico , Humanos , Mutación , Células Madre Neoplásicas/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Transducción de Señal

RESUMEN

Although tea catechins in green tea and green tea beverages must be stable to deliver good sensory quality and healthy benefits, they are always unstable during processing and storage. Ascorbic acid (AA) is often used to protect catechins in green tea beverages, and AA is easily oxidized to form dehydroascorbic acid (DHAA). However, the function of DHAA on the stability of catechins is not clear. The objective of this study was to determine the effects of DHAA on the stability of catechins and clarify the mechanism of effects by conducting a series of experiments that incubate DHAA with epigallocatechin gallate (EGCG) or catechins. Results showed that DHAA had a dual function on EGCG stability, protecting its stability by inhibiting hydrolysis and promoting EGCG consumption by forming ascorbyl adducts. DHAA also reacted with (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin (EGC) to form ascorbyl adducts, which destabilized them. After 9 h of reaction with DHAA, the depletion rates of EGCG, ECG, EC, and EGC were 30.08%, 22.78%, 21.45%, and 13.55%, respectively. The ability of DHAA to promote catechins depletion went from high to low: EGCG, ECG, EGC, and EC. The results are important for the processing and storage of tea and tea beverages, as well as the general exploration of synergistic functions of AA and catechins.

Asunto(s)

Catequina/química , Ácido Deshidroascórbico/química , Catequina/análogos & derivados , Ácido Deshidroascórbico/farmacología , Hidrólisis , Cinesis , Estructura Molecular , Oxidación-Reducción , Té/química , Temperatura

RESUMEN

Opioids and their antagonists alter vitamin C metabolism. Morphine binds to glutathione (l-γ-glutamyl-l-cysteinyl-glycine), an intracellular ascorbic acid recycling molecule with a wide range of additional activities. The morphine metabolite morphinone reacts with glutathione to form a covalent adduct that is then excreted in urine. Morphine also binds to adrenergic and histaminergic receptors in their extracellular loop regions, enhancing aminergic agonist activity. The first and second extracellular loops of adrenergic and histaminergic receptors are, like glutathione, characterized by the presence of cysteines and/or methionines, and recycle ascorbic acid with similar efficiency. Conversely, adrenergic drugs bind to extracellular loops of opioid receptors, enhancing their activity. These observations suggest functional interactions among opioids and amines, their receptors, and glutathione. We therefore explored the relative binding affinities of ascorbic acid, dehydroascorbic acid, opioid and adrenergic compounds, as well as various control compounds, to glutathione and glutathione-like peptides derived from the extracellular loop regions of the human beta 2-adrenergic, dopamine D1, histamine H1, and mu opioid receptors, as well as controls. Some cysteine-containing peptides derived from these receptors do bind ascorbic acid and/or dehydroascorbic acid and the same peptides generally bind opioid compounds. Glutathione binds not only morphine but also naloxone, methadone, and methionine enkephalin. Some adrenergic drugs also bind to glutathione and glutathione-like receptor regions. These sets of interactions provide a novel basis for understanding some ways that adrenergic, opioid and antioxidant systems interact during anesthesia and drug abuse and may have utility for understanding drug interactions.

Asunto(s)

Analgésicos Opioides/farmacología , Ácido Ascórbico/farmacología , Glutatión/metabolismo , Péptidos/farmacología , Ácido Deshidroascórbico/farmacología , Encefalina Metionina/farmacología , Humanos , Hidromorfona/análogos & derivados , Hidromorfona/orina , Metadona/farmacología , Morfina/farmacología , Naloxona/farmacología , Receptores Adrenérgicos beta 2/química , Receptores de Dopamina D1/química , Receptores Histamínicos H1/química , Receptores Opioides/química , Receptores Opioides mu/química

RESUMEN

Oxidative stress and inflammation are crucial factors that increase with age. In the progression of multiple age-related diseases, antioxidants and bioactive compounds have been recognized as useful antiaging agents. Oxidized or reduced vitamin C exerts different actions on tissues and has different metabolism and uptake. In this study, we analyzed the antiaging effect of vitamin C, both oxidized and reduced forms, in renal aging using laser microdissection, quantitative reverse-transcription polymerase chain reaction, and immunohistochemical analyses. In the kidneys of old SAM mice (10 months of age), a model of accelerated senescence, vitamin C, especially in the oxidized form (dehydroascorbic acid [DHA]) improves renal histology and function. Serum creatinine levels and microalbuminuria also decrease after treatment with a decline in azotemia. In addition, sodium-vitamin C cotransporter isoform 1 levels, which were increased during aging, are normalized. In contrast, the pattern of glucose transporter 1 expression is not affected by aging or vitamin C treatment. We conclude that oxidized and reduced vitamin C are potent antiaging therapies and that DHA reverses the kidney damage observed in senescence-accelerated prone mouse 8 to a greater degree.

Asunto(s)

Ácido Ascórbico/farmacología , Ácido Deshidroascórbico/farmacología , Inflamación/genética , Riñón/efectos de los fármacos , Transportadores de Sodio Acoplados a la Vitamina C/genética , Envejecimiento/genética , Envejecimiento/patología , Animales , Ácido Ascórbico/genética , Regulación de la Expresión Génica/efectos de los fármacos , Transportador de Glucosa de Tipo 1/genética , Humanos , Inflamación/patología , Riñón/ultraestructura , Ratones , Estrés Oxidativo/efectos de los fármacos

RESUMEN

Ascorbic acid better known as vitamin C, is a reducing carbohydrate needed for a variety of functions in the human body. The most important characteristic of ascorbic acid is the ability to donate two electrons, predestining it as a major player in balancing the physiological redox state and as a necessary cofactor in multiple enzymatic hydroxylation processes. Ascorbic acid can be reversibly oxidized in two steps, leading to semidehydroascorbic acid and dehydroascorbic acid, respectively. Further degradation is irreversible and generates highly reactive carbonyl-intermediates. These intermediates are able to induce glycation of proteins, a non-enzymatic and unspecific reaction of carbonyls with amino groups involved to several age-related diseases. In this study, we investigated the effect of ascorbic acid- and dehydroascorbic acid-induced glycation on PC12 cells, which represent a model for neuronal plasticity. We found that both applications of ascorbic acid or dehydroascorbic acid leads to glycation of cellular proteins, but that ascorbic acid interferes more with viability and neurite outgrowth compared with dehydroascorbic acid.

Asunto(s)

Ácido Ascórbico/farmacología , Proyección Neuronal/efectos de los fármacos , Animales , Supervivencia Celular/efectos de los fármacos , Ácido Deshidroascórbico/farmacología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Productos Finales de Glicación Avanzada/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células PC12 , Fosforilación/efectos de los fármacos , Ratas

RESUMEN

Zucchini (Cucurbita pepo subsp. pepo) is a seasonal vegetable with high nutritional and medical values. Many useful properties of this fruit are attributed to bioactive compounds. Zucchini fruits ("Yellow" and "Light Green" varieties) and four distinctive components (lutein, ß-carotene, zeaxanthin and dehydroascorbic acid) were selected. Firstly, the lutein, ß-carotene, zeaxanthin and dehydroascorbic acid contents were determined in these fruits. Then, in order to evaluate the safety and suitability of their use, different assays were carried out: (i) genotoxicity and anti-genotoxicity tests to determine the safety and DNA-protection against hydrogen peroxide; (ii) cytotoxicity; and (iii) DNA fragmentation and Annexin V/PI (Propidium Iodide) assays to evaluate the pro-apoptotic effect. Results showed that: (i) all the substances were non-genotoxic; (ii) all the substances were anti-genotoxic except the highest concentration of lutein; (iii) "Yellow" zucchini epicarp and mesocarp exhibited the highest cytotoxic activity (IC50 > 0.1 mg/mL and 0.2 mg/mL, respectively); and (iv) "Light Green" zucchini skin induced internucleosomal DNA fragmentation, ß-carotene being the possible molecule responsible for its pro-apoptotic activity. To sum up, zucchini fruit could play a positive role in human health and nutrition due to this fruit and its components were safe, able to inhibit significantly the H2O2-induced damage and exhibit anti-proliferative and pro-apoptotic activities toward HL60 (human promyelocytic leukemia cells) tumor cells. The information generated from this research should be considered when selecting potential accessions for breeding program purposes.

Asunto(s)

Cucurbita/química , Daño del ADN/efectos de los fármacos , Ácido Deshidroascórbico/farmacología , Luteína/farmacología , Zeaxantinas/farmacología , beta Caroteno/farmacología , Animales , Antineoplásicos Fitogénicos , Antioxidantes , Apoptosis/efectos de los fármacos , Fragmentación del ADN/efectos de los fármacos , Ácido Deshidroascórbico/análisis , Drosophila melanogaster/genética , Frutas/química , Células HL-60 , Promoción de la Salud , Humanos , Peróxido de Hidrógeno/farmacología , Luteína/análisis , Mutágenos , Valor Nutritivo , Fitoterapia , Zeaxantinas/análisis , beta Caroteno/análisis

RESUMEN

The high level of dehydroascorbic acid (DHA) in the lenticular tissue is an important risk factor for the development of age-related cataracts. In this study, the effects of DHA on structure and function of lens crystallins were studied in the presence of carnosine using gel mobility shift assay, different spectroscopic techniques, and lens culture analysis. The DHA-induced unfolding and aggregation of lens proteins were largely prevented by this endogenous dipeptide. The ability of carnosine to preserve native protein structure upon exposure to DHA suggests the essential role of this dipeptide in prevention of the senile cataract development. Although the DHA-modified α-crystallin was characterized by altered chaperone activity, functionality of this protein was significantly restored in the presence of carnosine. The increased proteolytic instability of DHA-modified lens proteins was also attenuated in the presence of carnosine. Furthermore, the assessment of lens culture suggested that DHA can induce significant lens opacity which can be prevented by carnosine. These observations can be explained by the pleiotropic functions of this endogenous and pharmaceutical compound, notably by its anti-glycation and anti-aggregation properties. In summary, our study suggests that carnosine may have therapeutic potential in preventing senile cataracts linked with the increased lenticular DHA generation, particularly under pathological conditions associated with the oxidative stress.

Asunto(s)

Carnosina/farmacología , Ácido Deshidroascórbico/antagonistas & inhibidores , Cristalino/efectos de los fármacos , alfa-Cristalinas/química , gamma-Cristalinas/química , Animales , Aspirina/farmacología , Catarata/metabolismo , Catarata/prevención & control , Bovinos , Ácido Deshidroascórbico/farmacología , Ensayo de Cambio de Movilidad Electroforética , Glicosilación/efectos de los fármacos , Cristalino/química , Cristalino/metabolismo , Sustancias Protectoras/farmacología , Técnicas de Cultivo de Tejidos

RESUMEN

INTRODUCTION: Accumulation of hypoxia inducible factor-1 alpha (HIF-1α) in malignant tissue is known to contribute to oncogenic progression and is inversely associated with patient survival. Ascorbic acid (AA) depletion in malignant tissue may contribute to aberrant normoxic activity of HIF-1α. While AA supplementation has been shown to attenuate HIF-1α function in malignant melanoma, the use of dehydroascorbic acid (DHA) as a therapeutic means to increase intracellular AA and modulate HIF-1α function is yet to be evaluated. Here we compared the ability of AA and DHA to increase intracellular vitamin C content and decrease the malignant potential of human melanoma by reducing the activity of HIF-1α. METHODS: HIF-1α protein accumulation was evaluated by western blot and transcriptional activity was evaluated by reporter gene assay using a HIF-1 HRE-luciferase plasmid. Protein expressions and subcellular localizations of vitamin C transporters were evaluated by western blot and confocal imaging. Intracellular vitamin C content following AA, ascorbate 2-phosphate (A2P), or DHA supplementation was determined using a vitamin C assay. Malignant potential was accessed using a 3D spheroid Matrigel invasion assay. Data was analyzed by One or Two-way ANOVA with Tukey's multiple comparisons test as appropriate with p<0.05 considered significant. RESULTS: Melanoma cells expressed both sodium dependent vitamin C (SVCT) and glucose (GLUT) transporters for AA and DHA transport respectively, however advanced melanomas responded favorably to AA, but not DHA. Physiological glucose conditions significantly impaired intracellular vitamin C accumulation following DHA treatment. Consequently, A2P and AA, but not DHA treated cells demonstrated lower HIF-1α protein expression and activity, and reduced malignant potential. The ability of AA to regulate HIF-1α was dependent on SVCT2 function and SVCT2 was not significantly inhibited at pH representative of the tumor microenvironment. CONCLUSIONS: The use of ascorbic acid as an adjuvant cancer therapy remains under investigated. While AA and A2P were capable of modulating HIF-1α protein accumulation/activity, DHA supplementation resulted in minimal intracellular vitamin C activity with decreased ability to inhibit HIF-1α activity and malignant potential in advanced melanoma. Restoring AA dependent regulation of HIF-1α in malignant cells may prove beneficial in reducing chemotherapy resistance and improving treatment outcomes.

Asunto(s)

Ácido Ascórbico/metabolismo , Ácido Ascórbico/farmacología , Ácido Deshidroascórbico/farmacología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Melanoma/tratamiento farmacológico , Melanoma/metabolismo , Ácido Ascórbico/análogos & derivados , Transporte Biológico/efectos de los fármacos , Línea Celular , Línea Celular Tumoral , Glucosa/metabolismo , Humanos , Melanoma/patología , Sodio/metabolismo , Transcripción Genética/efectos de los fármacos , Microambiente Tumoral/efectos de los fármacos

RESUMEN

Due to its anti-obesity effects, an adipocyte-derived hormone, leptin, has become important for the treatment of obesity. However, most obese subjects are in a state of leptin resistance, and endoplasmic reticulum (ER) stress is suggested to be involved in the pathophysiology of leptin resistance. Dehydroascorbic acid (DHAA), an oxidized form of vitamin C, was found to be increased in diabetes. In the present study, we investigated the possible effects of DHAA on the activation of ER stress and leptin resistance. A human neuroblastoma cell line, stably transfected with the Ob-Rb leptin receptor (SH-SY5Y-ObRb), was treated with DHAA. We found that DHAA upregulated ER stress-related genes such as GRP78, CHOP, and spliced XBP1. Moreover, leptin-induced STAT3 phosphorylation was hindered by DHAA. These results suggested that increases in the levels of DHAA might be harmful to neurons, contributing to defective leptin-responsive signaling.

Asunto(s)

Ácido Deshidroascórbico/farmacología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Retículo Endoplásmico/efectos de los fármacos , Leptina/farmacología , Neuronas/efectos de los fármacos , Línea Celular Tumoral , Retículo Endoplásmico/metabolismo , Chaperón BiP del Retículo Endoplásmico , Regulación de la Expresión Génica , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Leptina/genética , Leptina/metabolismo , Neuronas/metabolismo , Neuronas/patología , Oxidación-Reducción , Fosforilación/efectos de los fármacos , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismo , Respuesta de Proteína Desplegada/efectos de los fármacos , Proteína 1 de Unión a la X-Box/genética , Proteína 1 de Unión a la X-Box/metabolismo

RESUMEN

Ascorbic acid (AA), the reduced form of vitamin C, is incorporated into neurons via the sodium ascorbate co-transporter SVCT2. However, this transporter is not expressed in astrocytes, which take up the oxidized form of vitamin C, dehydroascorbic acid (DHA), via the facilitative hexose transporter GLUT1. Therefore, neuron and astrocyte interactions are thought to mediate vitamin C recycling in the nervous system. Although astrocytes are essential for the antioxidant defense of neurons under oxidative stress, a condition in which a large amount of ROS is generated that may favor the extracellular oxidation of AA and the subsequent neuronal uptake of DHA via GLUT3, potentially increasing oxidative stress in neurons. This study analyzed the effects of oxidative stress and DHA uptake on neuronal cell death in vitro. Different analyses revealed the presence of the DHA transporters GLUT1 and GLUT3 in Neuro2a and HN33.11 cells and in cortical neurons. Kinetic analyses confirmed that all cells analyzed in this study possess functional GLUTs that take up 2-deoxyglucose and DHA. Thus, DHA promotes the death of stressed neuronal cells, which is reversed by incubating the cells with cytochalasin B, an inhibitor of DHA uptake by GLUT1 and GLUT3. Additionally, the presence of glial cells (U87 and astrocytes), which promote DHA recycling, reverses the observed cell death of stressed neurons. Taken together, these results indicate that DHA promotes the death of stressed neurons and that astrocytes are essential for the antioxidative defense of neurons. Thus, the astrocyte-neuron interaction may function as an essential mechanism for vitamin C recycling, participating in the antioxidative defense of the brain.

Asunto(s)

Astrocitos/metabolismo , Ácido Deshidroascórbico/farmacología , Neuronas/patología , Estrés Oxidativo/efectos de los fármacos , Animales , Astrocitos/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Células Cultivadas , Corteza Cerebral/patología , Citocalasina B/farmacología , Desoxiglucosa/metabolismo , Femenino , Transportador de Glucosa de Tipo 1/metabolismo , Transportador de Glucosa de Tipo 3/metabolismo , Humanos , Cinética , Ratones , Modelos Biológicos , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuroprotección/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Ratas Sprague-Dawley

RESUMEN

Enhanced glucose requirement of cancer cells is associated with an increased glucose transport across plasma membrane that is mediated by a family of facilitated glucose transporter proteins, named GLUTs. GLUT1 is the main transporter in thyroid cancer cells. Glucose is the principal physiological substrate of GLUT1; however, it is also capable of transporting of oxidized form of vitamin C [i.e., dehydroascorbic acid (DHAA) which inside the cells is reduced to ascorbic acid (AA)]. The objective of this study was to determine the effect of normo-, hypo-, and hyperglycemia conditions on GLUT1-dependent intracellular ascorbate accumulation and viability of thyroid cancer cells. GLUT1 seems to be the main DHAA transporter in thyroid cancer cells because its knockdown by RNAi reduced DHAA accumulation by more than 80%. The results showed that in thyroid cancer cells high glucose inhibits both transport of AA and DHAA. Inhibition of vitamin C transport by glucose had a cytotoxic effect on the cells. However, stabilization of vitamin C in one of 2 forms (i.e., AA or DHAA) abolished this effect. These results suggest that cytotoxic effect is rather associated with extracellular accumulation of vitamin C and changes of its oxidation state than with intracellular level of ascorbate.

Asunto(s)

Ácido Ascórbico/metabolismo , Supervivencia Celular/efectos de los fármacos , Transportador de Glucosa de Tipo 1/fisiología , Glucosa/farmacología , Neoplasias de la Tiroides/metabolismo , Neoplasias de la Tiroides/patología , Ácido Ascórbico/farmacología , Transporte Biológico/efectos de los fármacos , Línea Celular Tumoral , Ácido Deshidroascórbico/farmacología , Regulación hacia Abajo , Expresión Génica/efectos de los fármacos , Transportador de Glucosa de Tipo 1/genética , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética

RESUMEN

Exposure of U937 cells to low concentrations of L-ascorbic acid (AA) is associated with a prompt cellular uptake and a further mitochondrial accumulation of the vitamin. Under the same conditions, dehydroascorbic acid (DHA) uptake was followed by rapid reduction and accumulation of identical intracellular levels of AA, however, in the absence of significant mitochondrial uptake. This event was instead observed after exposure to remarkably greater concentrations of DHA. Furthermore, experiments performed in isolated mitochondria revealed that DHA transport through hexose transporters and Na(+) -dependent transport of AA were very similar. These results suggest that the different subcellular compartmentalization of the vitamin is mediated by events promoting inhibition of mitochondrial AA transport, possibly triggered by low levels of DHA. We obtained results in line with this notion in intact cells, and more direct evidence in isolated mitochondria. This inhibitory effect was promptly reversible after DHA removal and comparable with that mediated by established inhibitors, as quercetin. The results presented collectively indicate that low intracellular concentrations of DHA, because of its rapid reduction back to AA, are a poor substrate for direct mitochondrial uptake. DHA concentrations, however, appear sufficiently high to mediate inhibition of mitochondrial transport of AA/DHA-derived AA.

Asunto(s)

Ácido Ascórbico/metabolismo , Transporte Biológico/efectos de los fármacos , Ácido Deshidroascórbico/farmacología , Mitocondrias/efectos de los fármacos , Transportadores de Sodio Acoplados a la Vitamina C/metabolismo , Línea Celular Tumoral , Humanos , Mitocondrias/metabolismo , Sodio/metabolismo , Células U937

RESUMEN

Arsenite directly triggers cytochrome c and Smac/DIABLO release in mitochondria isolated from U937 cells. These effects were not observed in mitochondria pre-exposed for 15 min to 10 µM L-ascorbic acid (AA). In other experiments, intact cells treated for 24-72 h with arsenite were found to die by apoptosis through a mechanism involving mitochondrial permeability transition. Pre-exposure (15 min) to low micromolar concentrations of AA and dehydroascorbic acid (DHA), resulting in identical cytosolic levels of the vitamin, had a diverse impact on cell survival, as cytoprotection was only observed after treatment with AA. Also the mitochondrial accumulation of the vitamin was restricted to AA exposure. An additional indication linking cytoprotection to the mitochondrial fraction of the vitamin was obtained in experiments measuring susceptibility to arsenite in parallel with loss of mitochondrial and cytosolic AA at different times after vitamin exposure. Finally, we took advantage of our recent findings that DHA potently inhibits AA transport to demonstrate that DHA abolishes all the protective effects of AA, under the same conditions in which the mitochondrial accumulation of the vitamin is prevented without affecting the overall cellular accumulation of the vitamin.

Asunto(s)

Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Arsenitos/antagonistas & inhibidores , Ácido Ascórbico/farmacología , Regulación Neoplásica de la Expresión Génica , Mitocondrias/efectos de los fármacos , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis , Arsenitos/toxicidad , Ácido Ascórbico/antagonistas & inhibidores , Transporte Biológico , Línea Celular Tumoral , Citocromos c/metabolismo , Citoprotección , Ácido Deshidroascórbico/farmacología , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Linfocitos/patología , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/genética , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Poro de Transición de la Permeabilidad Mitocondrial , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Transducción de Señal

RESUMEN

Water dispersible and biologically important molecule dehydroascorbic acid (DHA, capable to cross the blood brain barrier) coated Fe3O4 superparamagnetic nanoparticles having an average size of ∼6 nm were synthesized through one pot aqueous coprecipitation method under atmospheric air. An antioxidant ascorbic acid (AA) used in the synthesis oxidized itself to dehydroascorbic acid (DHA) to consume dissolved or available oxygen in reaction mixture which died away the oxidative impact of atmospheric air and formed DHA encapsulated the Fe3O4 nanoparticles which stabilized the Fe3O4 nanoparticles and significantly enhanced their colloidal solubility in water. Fe3O4 phase, superparamagnetic property, DHA coating and stable colloidal solubility in water were confirmed by means of XPS, VSM, IR and zeta potential analysis respectively. T1, T2 and T2(∗) weighted magnetic resonance imaging (MRI) and corresponding relaxivity (r1=0.416, r2=50.28 and r2(∗)=123.65 mM(-1) and r2/r1=120.86, r2(∗)r1=297.23) of colloidally dispersed DHA-coated nanoparticle water phantom revealed a strong contrast enhancement in T2 and T2(∗) weighted images. The compatibility of DHA-coated Fe3O4 nanoparticles toward human blood cells was examined by means of cell counting and cell morphological analysis with the use of optical microscope and scanning electron microscope imaging.

Asunto(s)

Células Sanguíneas/metabolismo , Materiales Biocompatibles Revestidos , Medios de Contraste , Ácido Deshidroascórbico , Compuestos Férricos , Nanopartículas de Magnetita/química , Ensayo de Materiales , Aire , Células Sanguíneas/ultraestructura , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Medios de Contraste/química , Medios de Contraste/farmacología , Ácido Deshidroascórbico/química , Ácido Deshidroascórbico/farmacología , Femenino , Compuestos Férricos/química , Compuestos Férricos/farmacología , Humanos , Imagen por Resonancia Magnética , Masculino

RESUMEN

The effects of L-ascorbic acid and its stable analogue L-ascorbic acid 2-glucoside on the restoration of liver mass and recovery of liver function after 70% partial hepatectomy (PH), were compared with other natural vitamin C analogues in rats in vivo. L-Ascorbic acid (100 mg/kg/d, intraperitoneally (i.p.))- and L-ascorbic acid 2-glucoside (50 mg/kg/d, i.p.)-treated rats showed an approximately 1.3-fold increase in the ratio of liver weight (LW) to body weight (BW), when compared to saline (as control)-, L-dehydroascorbic acid (150 mg/kg/d, i.p.)- and D-isoascorbic acid (150 mg/kg/d, i.p.)-administrated rats on day 3 after PH. Accordingly, 5-bromo-2-deoxyuridine-labeling index in the regenerating liver was significantly higher in L-ascorbic acid- and L-ascorbic acid 2-glucoside-treated rats compared with saline-, L-dehydroascorbic acid and D-isoascorbic acid-treated rats on day 1. In control rats, liver-related serum alanine aminotransferase (ALT) activity was rapidly elevated on day 1, and then decreased to near pre-operative levels on day 5 following PH. L-Ascorbic acid and L-ascorbic acid 2-glucoside significantly lowered the serum ALT on day 1 after PH compared with saline-, L-dehydroascorbic acid- and D-isoascorbic acid-administered rats. These results demonstrate that L-ascorbic acid and L-ascorbic acid 2-glucoside significantly promote the regeneration of liver mass and function with full recovery after liver injury.

Asunto(s)

Alanina Transaminasa/sangre , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/farmacología , Regeneración Hepática/efectos de los fármacos , Animales , Ácido Deshidroascórbico/farmacología , Hepatectomía , Ratas

RESUMEN

A U937 cell clone, in which low micromolar concentrations of ascorbic acid (AA) and dehydroascorbic acid (DHA) are taken up at identical rates, was used to investigate possible interactions between transport systems mediating cellular uptake of the two forms of the vitamin. Results obtained with different experimental approaches showed that DHA potently and reversibly inhibits AA uptake through Na(+)-AA cotransporters. Hence, a progressive increase in extracellular DHA concentrations in the presence of a fixed amount of AA caused an initial decrease in the net amount of vitamin C accumulated, and eventually, at higher levels, it caused an accumulation of the vitamin solely based on DHA uptake through hexose transporters. DHA-dependent inhibition of AA uptake was also detected in various other cell types. Taken together, our results provide evidence of a novel biological effect mediated by concentrations of DHA compatible with those produced at inflammatory sites.

Asunto(s)

Ácido Ascórbico/metabolismo , Ácido Deshidroascórbico/farmacología , Sodio/fisiología , Animales , Transporte Biológico Activo/efectos de los fármacos , Proteínas Portadoras/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Monocitos/efectos de los fármacos , Monocitos/metabolismo

RESUMEN

Recently, many clinical reports have suggested that the ascorbyl free radical (Asc(∙)) can be treated as a noninvasive, reliable, real-time marker of oxidative stress, but its generation mechanisms in human blood have rarely been discussed. In this study, we used upstream substances, enzyme inhibitors, and free radical scavengers to delineate the mechanisms of Asc(∙) formation in human platelet-rich plasma (PRP). Our results show that the doublet signal was detected in PRP samples by using electron spin resonance, and the hyperfine splitting of the doublet signal was a(H) = 1.88 gauss and g-factor = 2.00627, which was determined to be the Asc(∙). We observed that the inhibitors of NADPH oxidase (NOX), cyclooxygenase (COX), lipoxygenase (LOX), cytochrome P450 (CYP450), mitochondria complex III, and nitric oxide synthase (NOS), but not xanthine oxidase, diminished the intensity of the Asc(∙) signal dose dependently. All enzyme inhibitors showed no obvious antioxidant activity during a Fenton reaction assay. In summary, the obtained data suggest that Asc(∙) formation is associated with NOX, COX, LOX, CYP450, eNOS, and mitochondria in human PRP.

Asunto(s)

Ácido Deshidroascórbico/análogos & derivados , Plasma Rico en Plaquetas/metabolismo , Antioxidantes/metabolismo , Ácido Araquidónico/metabolismo , Inhibidores de la Ciclooxigenasa/farmacología , Inhibidores Enzimáticos del Citocromo P-450/farmacología , Sistema Enzimático del Citocromo P-450/metabolismo , Ácido Deshidroascórbico/sangre , Ácido Deshidroascórbico/farmacología , Espectroscopía de Resonancia por Spin del Electrón , Transporte de Electrón/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Radicales Libres/metabolismo , Humanos , Inhibidores de la Lipooxigenasa/farmacología , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Modelos Biológicos , NADPH Oxidasas/antagonistas & inhibidores , NADPH Oxidasas/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Superóxidos/metabolismo , Xantina Oxidasa/antagonistas & inhibidores , Xantina Oxidasa/metabolismo