Uptake of dehydroascorbic acid and ascorbic acid to isolated nerve terminals and secretory granules from ox neurohypophyses

When uptake of L-[14C]ascorbic acid ([14C]AA) to various organs in guinea-pigs was studied after intracardiac injection, the adenohypophysis, pars intermedia, and the neurohypophysis had an uptake per milligramme protein which was about half of the uptake to the adrenals. Adrenal uptake was 20 +/- 2.8 pmol mg-1 protein microCi-1 injected. The uptake to the different parts of the hypophysis was considerably higher than the uptake to pancreas, liver, kidney, spleen and other organs. When isolated nerve endings (neurosecretosomes) from ox neurohypophyses were incubated with a medium containing labelled dehydroascorbic acid ([14C]DHA), the uptake was much slower than when the medium contained labelled ascorbic acid. The uptake of [14C]DHA showed a linear dependence on concentration, and was not influenced by addition of Mg2+ and ATP. Addition of Mg2+ + ATP, omission of Ca2+ and Mg2+ or exchange of Na+ in the medium with K+ had no effect on the uptake of ascorbic acid. When isolated secretory granules from ox neurohypophyses were incubated with a medium containing [14C]DHA, uptake was considerably faster than the uptake when they were incubated in a medium containing [14C]AA. The uptake of dehydroascorbic acid was linear with the concentration in the medium and was not changed by addition of Mg2+ ATP. Addition of 10 mM NH4Cl or exchange of 120 mM K+ in the incubation medium with Na+ did not change the uptake of dehydroascorbic acid. The contents of copper, zinc, iron and cobalt were determined in isolated nerve endings (A) and membranes (B) as well as in lysate (C) from isolated neurosecretory granules. The results (in nmol mg-1 protein) were for Cu: (A): 0.25 +/- 0.01 (SEM), (B): 0.67 +/- 0.16, (C): 0.22 +/- 0.06; for Zn: (A): 0.53 +/- 0.13, (B): 6.97 +/- 0.75, (C): 1.8 +/- 0.53; and for Fe: (A): 15.6 +/- 1.9, (B): 6.92 +/- 0.32, (C): 3.15 +/- 0.43. In all preparations the cobalt content was below the detection limit (less than 5 pmol mg-1 protein).     

Ebselen is a dehydroascorbate reductase mimic, facilitating the recycling of ascorbate via mammalian thioredoxin systems

Ebselen is a selanazal drug recently revealed as a highly efficient peroxiredoxin mimic catalyzing the hydroperoxide reduction by the mammalian thioredoxin system [thioredoxin (Trx), thioredoxin reductase (TrxR), and NADPH]. The mammalian Trx system is a dehydroascorbic acid reductase recycling ascorbic acid essential for cell functions. Here we report that ebselen strongly facilitated the recycling of ascorbic acid by the TrxR both with and without Trx present. Reduction of dehydroascorbic acid by TrxR has a pH optimum of 6.4, and only approximately 55% of this activity at a physiological pH of 7.4. Ebselen at 6 microM enhances this reaction three-fold and with the same pH optimum of 6.4. The mechanism of the ebselen effect is suggested to involve reduction of dehydroascorbic acid by the ebselen selenol, a highly efficient two-electron reductant. Thus, ebselen acts as an antioxidant to lower the peroxide tone inside cells and to facilitate the recycling of dehydroascorbic acid to ascorbic acid, so as to increase the radical scavenging capacity of ascorbic acid directly or indirectly via vitamin E. The high ascorbic acid recycling efficiency of ebselen at pH 6.4 may play a major role in oxidatively stressed cells, where cytosol acidosis may trigger various responses, including apoptosis.     

Functions of vitamin C as a mediator of transmembrane electron transport in blood cells and related cell culture models

Vitamin C (ascorbic acid) is an important physiological antioxidant. Within cells, it is practically always present in the reduced form. Several enzymatic and nonenzymatic mechanisms have been reported to maintain this status. In the extracellular environment, oxidation of ascorbate leads to loss of vitamin because the oxidized form, dehydroascorbic acid, is unstable under physiological conditions. The intermediate ascorbate free radical, although rather long-lived for a free radical, quickly disproportionates into the two other forms, also leading to loss of vitamin. Protection from loss can only be achieved by cellular regeneration mechanisms, i.e., by uptake of dehydroascorbic acid and either storage or recycling, and by plasma-membrane mediated reduction of extracellular free radical or dehydroascorbic acid. Moreover, intracellular ascorbate can also serve as an electron donor for transmembrane reduction of external electron acceptors. However, the physiological significance of this function is as yet unknown. The results presented in the literature are sometimes conflicting as to the relative contributions of these different possibilities, which seem to differ in different cell types. In this short review, the various pathways of regeneration of ascorbate and their relative contributions to the avoidance of vitamin loss in plasma or cell culture medium are discussed.     

The excretion of ascorbic acid and its metabolites in uremia and hemodialysis

Renal excretion of ascorbic, dehydroascorbic and diketogulonic acids in uremia and relevant loss in hemodialysis are measured in comparison with those in patients with uremic syndrome (prior to hemodialysis) and in healthy subjects (control). Renal elimination of ascorbic acid was higher while of dehydroascorbic acid lower vs control. Elimination of diketogulonic acid was similar to control. In a session of hemodialysis, the organism loses 132.0 +/- 13.6 mg of ascorbic, 132.0 +/- 10.0 mg of dehydroascorbic and 204.0 +/- 9.0 mg of diketogulonic acid. 48-hour urinary losses of the patients reached 8.4 +/- 1.4, 19.6 +/- 1.1, 75.6 +/- 1.5 mg, respectively. Compared to control, hemodialysis patients lose the above acids 24.3, 2.7 and 4.6 times more.     

The contribution of ascorbic acid and dehydroascorbic acid to the protective role of pleura during inflammatory reactions

It is well-known that parapneumonic effusions lead to the formation of inflammatory exudates which contain an increasing amount of inflammatory cells, especially polymorphonuclear. At these pathological conditions characterized by oxidative stress, ascorbic acid (AA) plays an important role in quenching free radicals, so that it could protect neutrophils and mesothelial cells from oxidative damage. Besides that ascorbic acid and its metabolite dehydroascorbic acid (DHA) alters the sheep visceral and parietal pleura permeability. More specific ascorbic acid as well as dehydroascorbic acid decreases the permeability of pleura after addition on apical and basolateral side in both visceral and parietal pleurae. It seems that, AA and DHA have an opposite action upon pleura from that of the inflammatory mediators, like VEGF, which increases the permeability of pleura and causes mesothelial barrier dysfunction. The decrease of pleura permeability induced by AA and DHA suggest the hypothesis that AA and/or its metabolite DHA during inflammatory reactions not only protects mesothelial cells from oxidative damage, but also contributes to maintaining the mesothelial barrier function. Consequently, the inflammatory pleural fluid may be trapped in pleural space and the inflammation may be restricted, and have extension avoided.     

Inhibitory effect of ascorbic acid post-treatment on radiation-induced chromosomal damage in human lymphocytes in vitro

In the present study, the effect of exposure to ascorbic acid (vitamin C) after gamma-ray-induced chromosomal damage in cultured human lymphocytes was examined to explore the mechanism by which this antioxidant vitamin protects irradiated cells Non-irradiated lymphocytes were exposed to increasing concentrations of ascorbic acid (1-100 micro g/ml) and DNA damage was estimated using chromosomal aberration analysis and the comet assay. The results showed that ascorbic acid did not influence the frequency of chromosomal aberrations in non-irradiated cells, except at the highest concentration (20 micro g/ml), which induced breakage-type chromosomal aberrations. Vitamin C at the concentration of 50 micro g/ml caused DNA damage detected by the comet assay. A significant (34%) decrease in the frequency of chromosomal aberrations was observed in lymphocytes exposed to gamma-radiation and then cultured in the presence of ascorbic acid (1 micro g/ml). The removal of DNA breaks in cells exposed to 2 Gy of gamma-radiation was accelerated in the presence of ascorbic acid as determined by the comet assay, suggesting that it may stimulate DNA repair processes.     

Characteristics of ascorbic acid uptake by isolated ox neurohypophyseal nerve terminals and the influence of glucocorticoid and tri-iodothyronine on uptake

Isolated nerve endings (neurosecretosomes) from ox neurohypophyses took up L-[14C]ascorbic acid by a process or processes which showed energy dependence and which could be inhibited by unlabelled ascorbic acid in micromolar concentrations and by isoascorbic acid in millimolar concentrations, whereas dehydroascorbic acid only inhibited in concentrations of about 100 mM. The uptake showed saturation with increasing concentration of ascorbic acid and a Km value of 97 microM. Uptake was inhibited by increasing glucose concentration in the medium or by adding cytochalasin B, phloridzin, ethanol or probenecid to the medium. The uptake was inhibited by lowering the sodium concentration and by lack of calcium. These facts suggest the presence of both a glucose-dependent uptake and a sodium-dependent uptake. Cortisol and tri-iodothyronine inhibited uptake. This effect of cortisol, but not of tri-iodothyronine, was dependent on the presence of sodium in the medium. For both hormones it was still present when phloridzin or probenecid was added to the medium.     

Multifunctional PLGA particles containing poly(l-glutamic acid)-capped silver nanoparticles and ascorbic acid with simultaneous antioxidative and prolonged antimicrobial activity

A water-soluble antioxidant (ascorbic acid, vitamin C) was encapsulated together with poly(l-glutamic acid)-capped silver nanoparticles (AgNpPGA) within a poly(lactide-co-glycolide) (PLGA) polymeric matrix and their synergistic effects were studied. The PLGA/AgNpPGA/ascorbic acid particles synthesized by a physicochemical method with solvent/non-solvent systems are spherical, have a mean diameter of 775 nm and a narrow size distribution with a polydispersity index of 0.158. The encapsulation efficiency of AgNpPGA/ascorbic acid within PLGA was determined to be >90%. The entire amount of encapsulated ascorbic acid was released in 68 days, and the entire amount of AgNpPGAs was released in 87 days of degradation. The influence of PLGA/AgNpPGA/ascorbic acid on cell viability, generation of reactive oxygen species (ROS) in HepG2 cells, as well as antimicrobial activity against seven different pathogens was investigated. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay indicated good biocompatibility of these PLGA/AgNpPGA/ascorbic acid particles. We measured the kinetics of ROS formation in HepG2 cells by a DCFH-DA assay, and found that PLGA/AgNpPGA/ascorbic acid caused a significant decrease in DCF fluorescence intensity, which was 2-fold lower than that in control cells after a 5 h exposure. This indicates that the PLGA/AgNpPGA/ascorbic acid microspheres either act as scavengers of intracellular ROS and/or reduce their formation. Also, the results of antimicrobial activity of PLGA/AgNpPGA/ascorbic acid obtained by the broth microdilution method showed superior and extended activity of these particles. The samples were characterized using Fourier transform infrared spectroscopy, field-emission scanning electron microscopy, transmission electron microscopy, zeta potential and particle size analysis. This paper presents a new approach to the treatment of infection that at the same time offers a very pronounced antioxidant effect.     

Antiviral effects of ascorbic and dehydroascorbic acids in vitro

In the present study, ascorbic acid weakly inhibited the multiplication of viruses of three different families: herpes simplex virus type 1 (HSV-1), influenza virus type A and poliovirus type 1. Dehydroascorbic acid, an oxidized form of ascorbic acid and hence without reducing ability, showed much stronger antiviral activity than ascorbic acid, indicating that the antiviral activity of ascorbic acid is due to factors other than an antioxidant mechanism. Moreover, addition of 1 mM Fe3+, which oxidizes ascorbic acid to dehydroascorbic acid and also enhances the formation of hydroxyl radicals by ascorbic acid in the culture media, strongly enhanced the antiviral activity of ascorbic acid to a level significantly stronger than that of dehydroascorbic acid. Although both ascorbic acid and dehydroascorbic acid showed some cytotoxicity, the degree of cytotoxicity of the former was 10-fold higher than the latter, suggesting that the observed antiviral activity of ascorbic acid with and without ferric ion is, at least in part, a secondary result of the cytotoxic effect of the reagent, most likely due to the free radicals. However, the possibility that oxidation of ascorbic acid also contributed to the antiviral effects of ascorbic acid exists, in particular in the presence of ferric ion, since dehydroascorbic acid exhibited a very strong antiviral activity. Characterization of the mode of antiviral action of dehydroascorbic acid revealed that the addition of the reagent even at 11 h post infection almost completely inhibited the formation of progeny infectious virus in the infected cells, indicating that the reagent inhibits HSV-1 multiplication probably at the assembly process of progeny virus particles after the completion of viral DNA replication.     

Uptake of dehydroascorbic acid in high-GSH and normal-GSH dog erythrocytes

Uptake of dehydroascorbic acid (DHA) was studied in two types of dog erythrocytes with high GSH and normal GSH levels. Compared with ascorbic acid uptake, DHA produced a much greater ascorbic acid accumulation in dog erythrocytes. Both dog erythrocytes showed a concentration dependence of DHA uptake, and cellular ascorbic acid concentrations were significantly higher in high-GSH cells than in normal-GSH cells. Glucose and cytochalasin B inhibited DHA uptake. This suggests that DHA enters dog erythrocytes predominantly by the facilitated glucose transporter, particularly by the Glut 1 glucose transporter. The rate of glucose uptake was quite similar in the two types of cells. Compared with normal-GSH cells, high-GSH cells were more resistant to oxidative stress induced by high concentration of DHA. As a rapid entry of DHA inflicts on cells a heavy demand for GSH for its reduction to ascorbic acid, high-GSH cells containing a larger reserve of GSH have an advantage over normal-GSH cells in both ascorbic acid accumulation and resisting oxidative stress produced by DHA.     

Preparation and characterization of poly(D,L-lactide-co-glycolide) nanoparticles containing ascorbic acid

This paper is covering new, simplistic method of obtaining the system for controlled delivery of the ascorbic acid. Copolymer poly (D,L-lactide-co-glycolide) (DLPLG) nanoparticles are produced using physical method with solvent/nonsolvent systems where obtained solutions were centrifuged. The encapsulation of the ascorbic acid in the polymer matrix is performed by homogenization of water and organic phases. Particles of the DLPLG with the different content of ascorbic acid have different morphological characteristics, that is, variable degree of uniformity, agglomeration, sizes, and spherical shaping. Mean sizes of nanoparticles, which contain DLPLG/ascorbic acid in the ratio 85/150%, were between 130 to 200 nm depending on which stereological parameters are considered (maximal diameters Dmax, feret X, or feret Y). By introducing up to 15% of ascorbic acid, the spherical shape, size, and uniformity of DLPLG particles are preserved. The samples were characterized by infrared spectroscopy, scanning electron microscopy, stereological analysis, and ultraviolet spectroscopy.     

Simultaneous determination of inulin and p-aminohippuric acid in plasma and urine by reversed-phase high-performance liquid chromatography

A simple, accurate and sensitive high-performance liquid chromatographic method with UV detection was carried out to measure simultaneously plasma and urine concentrations of both p-aminohippuric acid and inulin. Following a simplified acid hydrolysis of the sample, the separation was carried out in 4 min using a C18 reversed-phase column with a flow-rate of 1 ml/min, and monitoring the absorbance at 280 nm. Within the investigated concentration ranges of inulin (0.1-3.2 mg/ml) and p-aminohippuric acid (0.0097-0.3 mg/ml), good linearity (r>0.99) was obtained. Within-run RSD ranged from 2.9 to 6.1% and between-run RSD ranged from 6.4 to 10%. Analytical recoveries were 101-112%, with little differences between plasma and urine samples. The detection limit was 1 microg/ml for all the analytes studied. This method might be ideal for renal function studies where a rapid and reproducible assessment of both renal glomerular filtration rate and blood flow-rate is required.     

Reduction of dehydroascrobic acid by aortic tissue of guinea pigs during acute radiation sickness

Summary Experiments were performed on guinea pigs. Acute radiation sickness provokes disturbance in the ability of the aortic tissue to reduce dehydroascrobic acid. In three days after irradiation this reducing faculty was halved and reassumed its normal value in 5 days.Diminution of the reducing faculty of aortic tissue with respect to dehydroascorbic acid in guinea pigs affected by inoizing radiation may serve as one of the causes of the drop of ascorbic acid content and of the sharp rise of the dehydroacetic acid concentration; this, in turn, affects the stability of the interstitial substance of connective tissue, and consequently the permeability and the fragility of the vascular wall.The rise of the intensity of dehydroascorbic acid reduction in five days after the irradiation, may evidently be regarded as a physiological protective reaction, connected with the increased ascorbic acid requirement.Presented by N. A. Kraevskii, Active Member, Acad. Med. Sci. USSR Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 51, No. 3, pp. 58–60, March, 1961     

The effect of vitamin C on antigen-induced bronchospasm.

The effect of vitamin C pretreatment in preventing ragweed-induced bronchospasm was evaluated in 6 ragweed-sensitive asthmatics studied in a double-blind randomized fashion. The patients received either lactose capsules or 500 mg of ascorbic acid and were studied out of season. Antigen dose-response curves were determined prior to the administration of lactose or ascorbic acid in each individual subject and subsequently after administration of ascorbic acid or lactose. Bothe PD20FEV1 (provocation dose necessary for a 20% reduction in forced expiratory volume in 1 second) and PD35SGaw (provocation dose necessary for a 35% reduction in specific airways conductance) were determined. In none of the six patients was there a change in baseline FEV 1 (p greater than 0.70) nor the overall average baseline specific airways conductance (rho greater than 0.90). Additionally, no statistically significant difference (p greater than 0.60) was noted between log PD35SGaw vitamin C day and lactose day. Likewise, no statistically significant difference (p greater than 0.60) was evident when comparing log PD20FEV1 lactose and ascorbic acid days. Vitamin C (500 MG) HAS NO PROTECTIVE EFFECT AGAINST RAGWEED ANTIGEN-INDUCED BRONCHOSPASM.     

Zum Wirkungsmechanismus von Vitamin C

Zusammenfassung In fast allen Vitamin-C-abhängigen Reaktionen wird Ascorbinsäure zu Dehydroascorbinsäure oxydiert. Die Wirkung des Vitamins ist in vitro nicht spezifisch, es kann durch andere Elektronendonatoren mit ähnlichen Eigenschaften aber ohne antiscorbutische Wirkung ersetzt werden. Trotzdem ist im Scorbut nur L-Ascorbinsäure (bzw. L-Dehydroascorbinsäure) imstande, die entsprechenden Symptome zu beheben. Diese Diskrepanz zwischen den in vitro- und in vivo-Versuchen wird auf die verhältnismäßig hohe Lipoidlöslichkeit der Dehydroascorbinsäure und die damit verbundenen besonders günstigen Verteilungseigenschaften im Organismus zurückgeführt.Der vergleichsweise großen Zahl Ascorbinsäure-oxydierender Prozesse stehen nur wenige Reaktionen in der tierischen Zelle gegenüber, die zu einer Reduktion des Vitamins führen. Diesen Vitamin C-erhaltenden Abläufen kommt damit besondere Bedeutung zu. Hier konnten eigene Untersuchungen einen Mechanismus nachweisen, nach dem aus Dehydroascorbinsäure und in der Zelle (in Spuren) vorliegender Ascorbinsäure durch Komproportionierung Semidehydroascorbinsäure-Radikal entsteht. Die radikalische Zwischenform des Vitamins wird in einer enzymatischen Reaktion zu Ascorbinsäure reduziert. Damit ist erstmalig auch für die tierische Zelle eine enzymatische Vitamin C-Reduktion gezeigt, die die verhältnismäßig zahlreichen Ascorbinsäure-oxydierenden Prozesse kompensiert.

---

Summary Ascorbic acid is oxidized to dehydroascorbic acid in almost all vitamin c-dependent reactions. The effect of this vitamin in vitro is not specific. It can be replaced by other electron donors with similar properties but without antiscorbutic action. In scurvy, however, onlyl-ascorbic acid (orl-dehydroascorbic acid, respectively) is able to cure the typical symptoms. This discrepancy between the results in vitro and in vivo can be explained by the relatively good lipid solubility of dehydroascorbic acid which results in an especially favourable distribution in the organism.While there occur comparatively many ascorbic acid oxidizing processes, there are only a few reactions in the animal cell leading to a reduction of the vitamin. Therefore, these vitamin c-preserving reactions are of particular significance. In this connexion studies in our laboratory gave evidence for the following mechanism: By comproportion of dehydroascorbic acid and ascorbic acid (present in trace amounts in the cell) a semidehydroascorbic acid radical is formed. This radical intermediate of the vitamin is reduced to ascorbic acid by an enzymatic reaction. Thereby for the first time an enzymatic reduction of vitamin c in the animal cell is shown compensating the relatively numerous ascorbic acid oxidizing reactions.

     

Ascorbic acid (vitamin C) modulates the mutagenic effects produced by an alkylating agent in vivo

Recent reports suggest that ascorbic acid (vitamin C) inhibits tumorigenesis as well as exerts a protective effect against mutagenesis in vitro; however, there is no information on its ability to affect gene mutations induced in vivo. In this study, we have investigated the antimutagenic effects of ascorbic acid on the frequency of 6-thioguanine-resistant (6-TGr) T-lymphocytes produced in Fischer 344 rats dosed with the direct-acting alkylating agent, N-ethyl-N-nitrosourea (ENU). The freqeuncy of 6-TG^r T-lymphocytes from the spleen measured five weeks after ENU treatment indicated that ENU produced a substantial mutagenic response. Pretreatment and/or post-treatment of rats with ascorbic acid administered in the drinking water appeared to inhibit the response, but the inhibition was statistically significant only when data from the various dosing schedules were pooled. In addition, there was no clear dose-dependency to the inhibitory effect of ascorbic acid. To further evaluate the time effects of the vitamin supplement on ENU mutagenicity, rats were exposed to the mutagen together with ascorbic acid, which was given continuously for the entire duration of the experiment. At specific times after ENU treatment, the frequency of 6-TG^r T-cells was determined in lymphocytes isolated from the spleen and the thymus. Time-dependent increases in the frequency of 6-TG^r T-cells were observed with ENU treatment; ascorbic acid significantly reduced the ENU-mediated mutagenic responses, most dramatically in the spleen at weeks 6 and 8 (P < 0.0001), and to a lesser extent in the thymus (P < 0.01 at week 6 and P < 0.006 at week 8). Our data suggest that ascorbic acid intake affects the in vivo mutagenicity of ENU, a direct-acting mutagen/carcinogen, and that the reported inhibitory effects of the antioxidant on carcinogenesis may be partially mediated by its effects on mutagenesis. Although it is difficult to extrapolate from rodent studies to humans, the results presented suggest an explanation for epidemiological data that link vitamin C ingestion with decreased cancer risk. © 1994 Wiley-Liss, Inc.     

Serum levels of vitamin C in young adults who chronically use drugs of abuse

Few studies have evaluated the relationship between drugs of abuse consumption and plasma levels of vitamin C. Because of the importance of vitamin C due to its role in prevention of acute and chronic diseases, this study was carried out with the purpose of testing the influence of consumption of drugs of abuse on the plasmatic levels of vitamin C (ascorbic acid) of 56 male chronic users of drugs of abuse with an age range of 16 to 40 years. The following was performed: 1) A survey of consumption to determine the kind, frequency and quantity of drug(s) used and 2) The plasmatic levels of vitamin C in fasting condition, using the Rue and Kuether method. The results obtained showed that 89% of the population under study used drugs for the first time before the age of 18, and 78.4% started with marijuana; 60.7% of them were mixed drug users. The average level of plasmatic ascorbic acid was 0.89 +/- 0.06 mg/dL, nevertheless, according to nutritional category, 76% have values greater than 0.4 mg/dL and 23.2% were at moderated risk (0.35 +/- 0.01 mg/dL) of vitamin C deficiency. Likewise, classifying them in the antioxidant category (according to the suggested values of Gey, 1993) it was obtained that 55.4% had suboptimal or inadequate concentrations to carry out its antioxidant protective function. The length of time of drug's consumption influenced on the ascorbic acid level too. It can be concluded that chronic consumption of drugs of abuse can negatively influence the plasmatic levels of ascorbic acid, leading these patients to a latent condition of vitamin C deficiency. Finally it is necessary to deepen the study of vitamin C levels of users of drugs of abuse.     

Dehydroascorbic acid for the treatment of acute ischemic stroke

In animal models of acute ischemic stroke, intravenous dehydroascorbic acid (DHAA), unlike ascorbic acid (AA), readily enters brain and is converted in both normal and ischemic brain into protective ascorbic acid. When given parenterally DHAA minimizes infarct volume and facilitates functional recovery. I hypothesize the same effect will occur in humans with acute ischemic stroke. Efficacy in reducing infarct volume is demonstrable in mice and rats even when DHAA is infused three hours after the experimental infarct. Moreover, there is fivefold mechanistic rational for DHA beside excellent pharmacokinetics and rapid penetration of brain and conversion to protective AA: (1) in ischemic brain, there is a precipitous decline in AA which can be reversed by intravenous DHAA; (2) after reduction of DHAA to AA in both normal and ischemic brain, AA can reduce oxidized vitamin E and glutathione, other protectors of brain against damaging reactive oxygen species which build up in ischemic brain; (3) AA itself can protect brain against damaging reactive oxygen species; (4) AA is an essential cofactor for several enzymes in brain including ten-eleven translocase-2 which upregulates production of protective molecules like brain-derived neurotrophic factor; and (5) DHAA after conversion to AA prevents both lipid oxidation and presumably oxidation of other labile substances (e.g., dopamine) in ischemic brain. In terms of safety, based on all available animal information, DHAA is safe in the proposed dosing regimen. For human clinical trials, the methodology for conducting the proposed animal safety, clinical pharmacology and phase II efficacy studies is straightforward. Finally, if DHAA preserved brain substance and function in humans, it could be employed in pre-hospital stroke patients.     

Identification and characterization of an ascorbic acid transporter in human granulosa-lutein cells.

Ascorbic acid serves a vital role as a pre-eminent antioxidant. In animals, it has been shown to be concentrated in granulosa and theca cells of the follicle, in luteal cells of the corpus luteum, and in the peripheral cytoplasm of the oocyte. We have previously identified hormonally-regulated ascorbic acid transporters in rat granulosa and luteal cells, and herein present preliminary evidence for the presence of a transporter for ascorbic acid in human granulosa-lutein cells. Granulosa-lutein cells were obtained from the follicular fluid of patients undergoing in-vitro fertilization. Following an overnight incubation, the cells were incubated with [14C]-ascorbic acid (0.15 microCi; 150 microM) and ascorbic acid uptake was determined. The uptake of ascorbic acid was saturable with a Michaeli's constant (Km) and maximum velocity (Vmax) of 21 microM and 3 pmol/10(6) cells/min respectively. Ouabain, low Na+ medium, and dinitrophenol significantly inhibited ascorbic acid uptake (P<0.05). Neither the presence of insulin, human chorionic gonadotrophin (HCG), insulin-like growth factor (IGF)-I, nor IGF-II affected the uptake of ascorbic acid in a statistically significant fashion. Following saturation of cellular uptake, the ascorbic acid level was estimated to be 1.04 pmoles/10(6) cells or approximately 1 mM, a high concentration similar to that seen in rat luteal cells. Active ascorbic acid transport in human granulosa-lutein cells appears to occur via a Na+ - and energy-dependent transporter, with high levels of ascorbic acid being accumulated in these cells.     

Changes in the ascorbic acid content of the white blood as an index of the vitamin C condition of the body

Summary In experiments on guinea pigs a relation has been established between the ascorbic acid content in the white cells and the dose of ascorbic acid administered. This permits us to conclude that the ascorbic acid content in the white cells may serve as a criterion of the vitamin C condition of the body.Presented by Active Member AMN SSSR B. A.