- Email: [email protected]
Ascorbic acid, myelination and associated disorders
PharmaNutrition 1 (2013) 98–100
Contents lists available at SciVerse ScienceDirect
PharmaNutrition journal homepage: www.elsevier.com/locate/phanu
Short communication
Ascorbic acid, myelination and associated disorders Fryad Rahman, Michel Fontes * Nutrition, Obesity and Thrombotic Risk (NORT), INSERM UMR 1062, Faculty of Medicine, University of Aix-Marseille, Marseille, France
A R T I C L E I N F O
A B S T R A C T
Article history: Available online 2 May 2013
Ascorbic acid has been considered, for a long time, only as an antioxidant. Since a few years, growing evidences, from literature, demonstrate that this molecule has other function. This is why different groups tried to find new targets for ascorbic acid, not only to prevent scurvy, but also as a drug. In this line of evidence, we report that high doses of ascorbic acid partially correct the phenotype of a Charcot– Marie–Tooth type 1A model, created in the lab. Based on this result, several clinical trials have been undertaken. Unfortunately, they have been controversial, primary outcomes never been reached, but tendencies observed. What conclusion could we draw? This will be discussed below. We published, in 2004, an article demonstrating that treatment of an animal model of CMT1A by high dose of ascorbic acid (AA), partially restores myelination and normal locomotion [1]. Our research was based on articles demonstrating that AA is necessary for myelination in co-culture of axon and Schwann cells [2–4]. This article was followed by clinical trials. Visioli et al. [5] recently published, in this journal, an article discussing AA treatment in Charcot–Marie–Tooth [5]. In this publication they use pharmacokinetics arguments to affirm that ascorbic acid could not be a treatment for CMT1A. In this paper, we will present arguments that demonstrate that this conclusion could be commented, as numerous arguments, based on literature, are not present in the article. We will thus expose these additional arguments, in the following sections. ß 2013 Elsevier B.V. All rights reserved.
Keywords: Ascorbic acid Myelin Charcot–Marie–Tooth
1. AA concentration in blood and in tissues Most of pharmacokinetics studies, although being rare, are postprandial studies, evaluating blood concentration a few hours after AA intake. Before CMT clinical trials, we did not have a clear view on long term blood concentration after long term supplementation with AA. Only one paper [6] provides information on this point. Supplementation is well documented up to 1 g/day but not using higher concentration. However, from data published from clinical trials [7,8], it is clear that supplementation with high doses led to an increase in concentration of AA, proportional to the dose administrated to patients (see Table 3 of Micallef et al.). As techniques used to evaluate AA blood concentration, it is difficult to conclude. However, we could evaluate the percentage (%) of increase in different trials. In the Italian/English trial [8], supplementation with 1.5 g/day results in increasing blood concentration by 33% (first chapter, p. 324). In our trial, supplementation by 1 g/day results in 40% increase as supplementation by 3 g/day results in 51% increase in blood concentration. We will comment about the potential impact of these data in the
* Corresponding author at: Nutrition, Obesity and Thrombotic Risk (NORT), INSERM UMR 1062, Faculty of Medicine, University of Aix-Marseille, 13385 Marseille cedex 5, France. Tel.: +33 (0)4 91 32 44 30; fax: +33 (0)4 91 32 43 87. E-mail address: [email protected] (M. Fontes). 2213-4344/$ – see front matter ß 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.phanu.2013.04.003
following section, but we could observe that blood concentration of AA in the I/E trial never reach the concentration of 1 g supplementation in our trial, a concentration that we demonstrate having no effect on CMT phenotype of patients. In addition, we will point out that AA short-term kinetics did not reflect long term ‘‘at equilibrium’’ concentration that is clearly increased when you increase AA supplementation. What could be the mechanisms of these differences? From what we know from AA transport, two major players, apart from DHA transport, are involved [9,10]. One transporter, SVCT1, allow entrance of AA in circulation as well as re-intakes by kidney, and the second, SVCT2, is involved in transport inside cell. We will discuss this in the following section.
2. AA and transporter/receptor SVCT1, the protein that transport AA from gut to blood circulation, exists as different variant forms [11,12]. About 10/ 20% of the population has a poor allele (20% of normal allele) [12]. As a consequence these persons have a low concentration of AA in blood [13]. As we point above, is it why we have good and poor responders in trials? This could be the case in the population studied by Burns et al. [14] we should thus do not consider that a cohort of patient is homogeneous, but is composed of persons that could have different response to AA administration.
F. Rahman, M. Fontes / PharmaNutrition 1 (2013) 98–100
Looking carefully at our clinical trial report, we could observe two events. First, providing more AA led to a steady state of AA concentration higher (average of 86 mM for 1 g/group and 106 mM for 3 g/group). Moreover, we could see that standard deviation is high, and that a concentration higher than 160 mM could be achieved in some patients. This is why we should not consider averages but statistic distribution. Genetic background is important, variants of SVCT1 being probably an important part of the story. An important question is which role the different molecule binding AA plays in cell differentiation. SVCT2 is a transmembrane protein allowing transporting AA inside cell, but the same molecule could also play the role of a receptor, as it is highly specific of AA. No variant of SVCT2 has been described so far, and this one of the most conserved protein in evolution (70% identity between human and Drosophila). There is thus a high evolutionary pressure on this molecule. We may note that gradient in retinoic acid is not caused by differentiation distribution in circulation but by differential expression of the receptor in different part of the body. This is why intracellular concentration should be considered and not only concentration in blood. Our recent findings suggest that signalling through SVCT2 is a key event, not only for myelination. From our recent results it seems to be the same for ascorbic acid and our unpublished data suggest that AA is a key signalling molecule in embryology and development as retinoic acid is (Rahman et al., unpublished). 3. Which conclusions could we draw from CMT clinical trials? We will comment the two first clinical trials. The paper by Burns et al. [14] shows (Fig. 3B of the paper) that, at least, a sub population of patients take advantage of ascorbic acid treatment. Unfortunately, p value was only 0.06, a value so close to significantly! However, these data clearly establish that there are at least two sub types of patients, poor and good responders. Regarding our trial [7], if statistical significativity has not been obtained using the CMTNS score, although a tendency was observed, statistical significance is obtained using another score, a clinical score, CMTES. Unfortunately, this score was not the primary outcome otherwise this trial has been positive. In addition, you could see (Fig. 2B of Micallef et al.) that treatment with either a placebo or 1 g/day did not present any differences on the score. On the contrary, 3 g/day treatment stops progression of the disorder vs placebo or 1 g (p value = 0.02). This clearly demonstrates that increasing doses are more efficient. However, these data results from a post hoc analysis. Finally, the Italian/English trial deduced, from their clinical trial, that AA treatment has no benefit for patients. However, as we point above, blood concentration of AA did not reach concentration that seems to have an effect, using CMTES, in our trial. Looking to our previous models of CMT1A [15,16], as well as in published in vitro experiments, Schwann cells seems to accept PMP22 over expression up to a threshold about 80% over expression. If we consider the level of expression of PMP22 that should be repressed to obtain an effect by AA treatment, we obtain a blood concentration of between 100/150 mM. Obviously, these data are just an evaluation, but they could be reached [17] by a continuous treatment of more than 2 g/day. A final question, is the evaluation score, CMTNS appropriate to monitor the progress of a slightly evolving disorder as CMT1A? It is clear from our study that this is not the case, likely because NCV is not related to severity and evolution of the disease. Building a new grid of evaluation will probably be a major task in the future, in order to correctly evaluate the impact of a treatment for CMT1A. This is part of a discussion pointing that big effort have been done on molecular mechanisms of slightly evolving genetic disorders
99
but only a few regarding natural history of these diseases. Exchanges between scientists and clinicians will probably be strengthened in the future. Finally, just a comment, when we say that we could see difference in concentration in animals did not mean that it exists only that we did not a good technique to test it in tissue (it is not easy). 4. AA and myelination It has been described, since 1986 (see above), that AA is necessary for in vitro myelination in Schwann cell/axon co-culture. AA is acting not through its antioxidant activity but probably through other mechanisms involving regulation of the cAMP pool [18]. It is likely that the new function of AA we report, a weak inhibitor of adenylate cyclase [19], could be involved. It is interesting to note that a new GPCR protein has been described as a key factor in myelination [20]. This protein is closely associated (physically) to adenylate cyclase and the phenotype of animals invalidated for this protein is saved by forskolin, elevating the cAMP level. A first paper of Gess et al. [21], confirm that AA has an impact in PMP22 expression. Moreover, a more recent paper of the same group [22] demonstrates the impact of AA in myelin differentiation. They use heterozygotes mouse, invalidated for SVCT2 [23], a receptor/ transporter of AA. In these animals, production of AA is normal but AA could not enter cells. These heterozygote animals (homozygotes are lethal) present a demyelinating neuropathy similar to CMT1A. In addition, it confirms that dosages of AA inside cells are crucial for myelination. This paper is crucial regarding the thematic. These data clearly demonstrate that AA play a role in myelination, probably as a global modulator of the intracellular cAMP pool, via its action on adenylate cyclases. 5. Conclusion AA has been for long only considered as a vitamin, and a lot of discussions regarding RDA have been initiated. However, if we consider AA as a key molecule, and not only an antioxidant molecule, and, may be as a drug, we could not consider the same type of arguments that lead to RDA. This is why recommendation as well as all consideration, coming from nutrition (the vitamin concept) is probably not appropriate to explain cell signalling and cell differentiation linked to ascorbic acid. Regarding the impact on CMT patients, AA has been approved up to 4 g/day by regulatory agencies. Moreover, a recent paper of Levine and al, demonstrate that high blood concentration of AA, obtained by IV, and did not present any safety problem [24]. So, why not recommend to CMT1A patients to take 3 g/day, a concentration approved by agencies and that has been demonstrated to be safe? If this open assay take place, it will be interesting to evaluate degradation of CMT1A sign in this patients, versus untreated. We may note that AA did not restore a normal locomotion, but seems to stop progression. Therefore, what will be the evolution of the disorder if asymptomatic patients are treated in early phases? A last comment, treatment of disorder during all the life should be only envisaged using safe drugs. AA is clearly one of them. Conflict of interest Aix-Marseille University, INSERM, and AFM hold a patent on ascorbic acid for CMT1A treatment. References [1] Passage E, Norreel JC, Noack-Fraissignes P, Sanguedolce V, Pizant J, Thirion X, et al. Ascorbic acid treatment corrects the phenotype of a mouse model of Charcot–Marie–Tooth disease. Nature Medicine 2004;10:396–401.
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F. Rahman, M. Fontes / PharmaNutrition 1 (2013) 98–100
[2] Carey DJ, Todd MS. Schwann cell myelination in a chemically defined medium: demonstration of a requirement for additives that promote Schwann cell extracellular matrix formation. Brain Research 1987;429(1):95–102. [3] Eldridge CF, Bunge MB, Bunge RP, Wood PM. Differentiation of axon-related Schwann cells in vitro ascorbic acid regulates basal lamina assembly and myelin formation. Journal of Cell Biology 1987;105(2):1023–34. [4] Plant GW, Currier PF, Cuervo EP, Bates ML, Pressman Y, Bunge MB, et al. Purified adult ensheathing glia fail to myelinate axons under culture conditions that enable Schwann cells to form myelin. Journal of Neuroscience 2002;22(14):6083–91. [5] Visioli F, Reilly MM, Rimoldi M, Solari A, Pareysonc D, CMT-TRIAAL & CMTTRAUK Groups. Vitamin C and Charcot–Marie–Tooth 1A: pharmacokinetic considerations. PharmaNutrition 2013;1:10–2. [6] Levine M, Conry-Cantilena C, Wang Y, Welch RW, Washko PW, Dhariwal KR, et al. Vitamin C pharmacokinetics in healthy volunteers: evidence for a recommended dietary allowance. Proceedings of the National Academy of Sciences of the United States of America 1996;93:3704–9. [7] Micallef J, Attarian S, Dubourg O, Gonnaud PM, Hogrel JY, Stojkovic T, et al. Effect of ascorbic acid in patients with Charcot–Marie–Tooth disease type 1A: a multicentre, randomised, double-blind, placebo-controlled trial. Lancet Neurology 2009;8:1103–10. [8] Pareyson D, Reilly MM, Schenone A, Fabrizi GM, Cavallaro T, Santoro L, et al. Ascorbic acid in Charcot–Marie–Tooth disease type 1A (CMT-TRIAAL and CMTTRAUK): a double-blind randomised trial. Lancet Neurology 2011;10:320–8. [9] Tsukaguchi H, Tokui T, Mackenzie B, Berger UV, Chen XZ, Wang Y, et al. A family of mammalian Na+-dependent L-ascorbic acid transporters. Nature 1999;399:70–5. [10] Daruwala R, Song J, Koh WS, Rumsey SC, Levine M. Cloning and functional characterization of the human sodium-dependent AA transporters hSVCT1 and hSVCT2. FEBS Letters 1999;460(3):480–4. [11] Eck P, Erichsen HC, Taylor JG, Yeager M, Hughes AL, Levine M, et al. Comparison of the genomic structure and variation in the two human sodium-dependent vitamin C transporters, SLC23A1 and SLC23A2. Human Genetics 2004;115: 285–94. [12] Corpe CP, Tu H, Eck P, Wang J, Faulhaber-Walter R, Schnermann J, et al. Vitamin C transporter Slc23a1 links renal reabsorption, vitamin C tissue accumulation, and perinatal survival in mice. Journal of Clinical Investigation 2010;120(4):1069–83.
[13] Timpson NJ, Forouhi NG, Brion MJ, Harbord RM, Cook DG, Johnson P, et al. Genetic variation at the SLC23A1 locus is associated with circulating concentrations of L-ascorbic acid (vitamin C): evidence from 5 independent studies with >15,000 participants. American Journal of Clinical Nutrition 2010;92(2):375–82. [14] Burns J, Ouvrier RA, Yiu EM, Joseph PD, Kornberg AJ, Fahey MC, et al. Ascorbic acid for Charcot–Marie–Tooth disease type 1A in children: a randomised, double-blind, placebo-controlled, safety and efficacy trial. Lancet Neurology 2009;8:537–44. [15] Huxley C, Passage E, Manson A, Putzu G, Figarella-Branger D, Pellissier JF, et al. Construction of a mouse model of the CMT1A disease by YAC injection in the murine oocyte. Human Molecular Genetics 1996;5:563–9. [16] Huxley C, Passage E, Robertson AM, Youl B, Huston S, Manson A, et al. Correlation between varying levels of PMP22 expression and the degree of demyelination and reduction in verve conduction velocity in transgenic mice. Human Molecular Genetics 1998;7:449–58. [17] Padayatty SJ, Sun H, Wang Y, Riordan HD, Hewitt SM, Katz A, et al. Vitamin C pharmacokinetics: implications for oral and intravenous use. Annals of Internal Medicine 2004;140:533–7. [18] Kaya F, Belin S, Bourgeois P, Micaleff J, Blin O, Fontes M. Ascorbic acid inhibits PMP22 expression by reducing cAMP levels. Neuromuscular Disorders 2007;17:248–53. [19] Kaya F, Belin S, Diamantidis G, Fontes M. Ascorbic acid is a regulator of the intracellular cAMP concentration: old molecule, new functions? FEBS Letters 2008;582(25–26):3614–8. [20] Monk KR, Naylor SG, Glenn TD, Mercurio S, Perlin JR, Dominguez C, et al. A G protein-coupled receptor is essential for Schwann cells to initiate myelination. Science 2009;325:1402–5. [21] Gess B, Lohmann C, Halfter H, Young P. Sodium-dependent vitamin C transporter 2 (SVCT2) is necessary for the uptake of L-ascorbic acid into Schwann cells. Glia 2010;58:287–99. [22] Gess B, Rohr D, Fledrich R, Sereda MW, Kleffner I, Humberg A, et al. Transporter 2 deficiency causes hypomyelination and extracellular matrix defects in the peripheral nervous system. Journal of Neuroscience 2011;31(47):17180–92. [23] Sotiriou S, Gispert S, Cheng J, Wang Y, Chen A, Hoogstraten-Miller S, et al. Ascorbic-acid transporter Slc23a1 is essential for vitamin C transport into the brain and for perinatal survival. Nature Medicine 2002;8:514–7. [24] Padayatty SJ, Sun AY, Chen Q, Espey MG, Drisko J, Levine M, et al. intravenous use by complementary and alternative medicine practitioners and adverse effects. PLoS ONE 2010;5:e11414.
Contents lists available at SciVerse ScienceDirect
PharmaNutrition journal homepage: www.elsevier.com/locate/phanu
Short communication
Ascorbic acid, myelination and associated disorders Fryad Rahman, Michel Fontes * Nutrition, Obesity and Thrombotic Risk (NORT), INSERM UMR 1062, Faculty of Medicine, University of Aix-Marseille, Marseille, France
A R T I C L E I N F O
A B S T R A C T
Article history: Available online 2 May 2013
Ascorbic acid has been considered, for a long time, only as an antioxidant. Since a few years, growing evidences, from literature, demonstrate that this molecule has other function. This is why different groups tried to find new targets for ascorbic acid, not only to prevent scurvy, but also as a drug. In this line of evidence, we report that high doses of ascorbic acid partially correct the phenotype of a Charcot– Marie–Tooth type 1A model, created in the lab. Based on this result, several clinical trials have been undertaken. Unfortunately, they have been controversial, primary outcomes never been reached, but tendencies observed. What conclusion could we draw? This will be discussed below. We published, in 2004, an article demonstrating that treatment of an animal model of CMT1A by high dose of ascorbic acid (AA), partially restores myelination and normal locomotion [1]. Our research was based on articles demonstrating that AA is necessary for myelination in co-culture of axon and Schwann cells [2–4]. This article was followed by clinical trials. Visioli et al. [5] recently published, in this journal, an article discussing AA treatment in Charcot–Marie–Tooth [5]. In this publication they use pharmacokinetics arguments to affirm that ascorbic acid could not be a treatment for CMT1A. In this paper, we will present arguments that demonstrate that this conclusion could be commented, as numerous arguments, based on literature, are not present in the article. We will thus expose these additional arguments, in the following sections. ß 2013 Elsevier B.V. All rights reserved.
Keywords: Ascorbic acid Myelin Charcot–Marie–Tooth
1. AA concentration in blood and in tissues Most of pharmacokinetics studies, although being rare, are postprandial studies, evaluating blood concentration a few hours after AA intake. Before CMT clinical trials, we did not have a clear view on long term blood concentration after long term supplementation with AA. Only one paper [6] provides information on this point. Supplementation is well documented up to 1 g/day but not using higher concentration. However, from data published from clinical trials [7,8], it is clear that supplementation with high doses led to an increase in concentration of AA, proportional to the dose administrated to patients (see Table 3 of Micallef et al.). As techniques used to evaluate AA blood concentration, it is difficult to conclude. However, we could evaluate the percentage (%) of increase in different trials. In the Italian/English trial [8], supplementation with 1.5 g/day results in increasing blood concentration by 33% (first chapter, p. 324). In our trial, supplementation by 1 g/day results in 40% increase as supplementation by 3 g/day results in 51% increase in blood concentration. We will comment about the potential impact of these data in the
* Corresponding author at: Nutrition, Obesity and Thrombotic Risk (NORT), INSERM UMR 1062, Faculty of Medicine, University of Aix-Marseille, 13385 Marseille cedex 5, France. Tel.: +33 (0)4 91 32 44 30; fax: +33 (0)4 91 32 43 87. E-mail address: [email protected] (M. Fontes). 2213-4344/$ – see front matter ß 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.phanu.2013.04.003
following section, but we could observe that blood concentration of AA in the I/E trial never reach the concentration of 1 g supplementation in our trial, a concentration that we demonstrate having no effect on CMT phenotype of patients. In addition, we will point out that AA short-term kinetics did not reflect long term ‘‘at equilibrium’’ concentration that is clearly increased when you increase AA supplementation. What could be the mechanisms of these differences? From what we know from AA transport, two major players, apart from DHA transport, are involved [9,10]. One transporter, SVCT1, allow entrance of AA in circulation as well as re-intakes by kidney, and the second, SVCT2, is involved in transport inside cell. We will discuss this in the following section.
2. AA and transporter/receptor SVCT1, the protein that transport AA from gut to blood circulation, exists as different variant forms [11,12]. About 10/ 20% of the population has a poor allele (20% of normal allele) [12]. As a consequence these persons have a low concentration of AA in blood [13]. As we point above, is it why we have good and poor responders in trials? This could be the case in the population studied by Burns et al. [14] we should thus do not consider that a cohort of patient is homogeneous, but is composed of persons that could have different response to AA administration.
F. Rahman, M. Fontes / PharmaNutrition 1 (2013) 98–100
Looking carefully at our clinical trial report, we could observe two events. First, providing more AA led to a steady state of AA concentration higher (average of 86 mM for 1 g/group and 106 mM for 3 g/group). Moreover, we could see that standard deviation is high, and that a concentration higher than 160 mM could be achieved in some patients. This is why we should not consider averages but statistic distribution. Genetic background is important, variants of SVCT1 being probably an important part of the story. An important question is which role the different molecule binding AA plays in cell differentiation. SVCT2 is a transmembrane protein allowing transporting AA inside cell, but the same molecule could also play the role of a receptor, as it is highly specific of AA. No variant of SVCT2 has been described so far, and this one of the most conserved protein in evolution (70% identity between human and Drosophila). There is thus a high evolutionary pressure on this molecule. We may note that gradient in retinoic acid is not caused by differentiation distribution in circulation but by differential expression of the receptor in different part of the body. This is why intracellular concentration should be considered and not only concentration in blood. Our recent findings suggest that signalling through SVCT2 is a key event, not only for myelination. From our recent results it seems to be the same for ascorbic acid and our unpublished data suggest that AA is a key signalling molecule in embryology and development as retinoic acid is (Rahman et al., unpublished). 3. Which conclusions could we draw from CMT clinical trials? We will comment the two first clinical trials. The paper by Burns et al. [14] shows (Fig. 3B of the paper) that, at least, a sub population of patients take advantage of ascorbic acid treatment. Unfortunately, p value was only 0.06, a value so close to significantly! However, these data clearly establish that there are at least two sub types of patients, poor and good responders. Regarding our trial [7], if statistical significativity has not been obtained using the CMTNS score, although a tendency was observed, statistical significance is obtained using another score, a clinical score, CMTES. Unfortunately, this score was not the primary outcome otherwise this trial has been positive. In addition, you could see (Fig. 2B of Micallef et al.) that treatment with either a placebo or 1 g/day did not present any differences on the score. On the contrary, 3 g/day treatment stops progression of the disorder vs placebo or 1 g (p value = 0.02). This clearly demonstrates that increasing doses are more efficient. However, these data results from a post hoc analysis. Finally, the Italian/English trial deduced, from their clinical trial, that AA treatment has no benefit for patients. However, as we point above, blood concentration of AA did not reach concentration that seems to have an effect, using CMTES, in our trial. Looking to our previous models of CMT1A [15,16], as well as in published in vitro experiments, Schwann cells seems to accept PMP22 over expression up to a threshold about 80% over expression. If we consider the level of expression of PMP22 that should be repressed to obtain an effect by AA treatment, we obtain a blood concentration of between 100/150 mM. Obviously, these data are just an evaluation, but they could be reached [17] by a continuous treatment of more than 2 g/day. A final question, is the evaluation score, CMTNS appropriate to monitor the progress of a slightly evolving disorder as CMT1A? It is clear from our study that this is not the case, likely because NCV is not related to severity and evolution of the disease. Building a new grid of evaluation will probably be a major task in the future, in order to correctly evaluate the impact of a treatment for CMT1A. This is part of a discussion pointing that big effort have been done on molecular mechanisms of slightly evolving genetic disorders
99
but only a few regarding natural history of these diseases. Exchanges between scientists and clinicians will probably be strengthened in the future. Finally, just a comment, when we say that we could see difference in concentration in animals did not mean that it exists only that we did not a good technique to test it in tissue (it is not easy). 4. AA and myelination It has been described, since 1986 (see above), that AA is necessary for in vitro myelination in Schwann cell/axon co-culture. AA is acting not through its antioxidant activity but probably through other mechanisms involving regulation of the cAMP pool [18]. It is likely that the new function of AA we report, a weak inhibitor of adenylate cyclase [19], could be involved. It is interesting to note that a new GPCR protein has been described as a key factor in myelination [20]. This protein is closely associated (physically) to adenylate cyclase and the phenotype of animals invalidated for this protein is saved by forskolin, elevating the cAMP level. A first paper of Gess et al. [21], confirm that AA has an impact in PMP22 expression. Moreover, a more recent paper of the same group [22] demonstrates the impact of AA in myelin differentiation. They use heterozygotes mouse, invalidated for SVCT2 [23], a receptor/ transporter of AA. In these animals, production of AA is normal but AA could not enter cells. These heterozygote animals (homozygotes are lethal) present a demyelinating neuropathy similar to CMT1A. In addition, it confirms that dosages of AA inside cells are crucial for myelination. This paper is crucial regarding the thematic. These data clearly demonstrate that AA play a role in myelination, probably as a global modulator of the intracellular cAMP pool, via its action on adenylate cyclases. 5. Conclusion AA has been for long only considered as a vitamin, and a lot of discussions regarding RDA have been initiated. However, if we consider AA as a key molecule, and not only an antioxidant molecule, and, may be as a drug, we could not consider the same type of arguments that lead to RDA. This is why recommendation as well as all consideration, coming from nutrition (the vitamin concept) is probably not appropriate to explain cell signalling and cell differentiation linked to ascorbic acid. Regarding the impact on CMT patients, AA has been approved up to 4 g/day by regulatory agencies. Moreover, a recent paper of Levine and al, demonstrate that high blood concentration of AA, obtained by IV, and did not present any safety problem [24]. So, why not recommend to CMT1A patients to take 3 g/day, a concentration approved by agencies and that has been demonstrated to be safe? If this open assay take place, it will be interesting to evaluate degradation of CMT1A sign in this patients, versus untreated. We may note that AA did not restore a normal locomotion, but seems to stop progression. Therefore, what will be the evolution of the disorder if asymptomatic patients are treated in early phases? A last comment, treatment of disorder during all the life should be only envisaged using safe drugs. AA is clearly one of them. Conflict of interest Aix-Marseille University, INSERM, and AFM hold a patent on ascorbic acid for CMT1A treatment. References [1] Passage E, Norreel JC, Noack-Fraissignes P, Sanguedolce V, Pizant J, Thirion X, et al. Ascorbic acid treatment corrects the phenotype of a mouse model of Charcot–Marie–Tooth disease. Nature Medicine 2004;10:396–401.
100
F. Rahman, M. Fontes / PharmaNutrition 1 (2013) 98–100
[2] Carey DJ, Todd MS. Schwann cell myelination in a chemically defined medium: demonstration of a requirement for additives that promote Schwann cell extracellular matrix formation. Brain Research 1987;429(1):95–102. [3] Eldridge CF, Bunge MB, Bunge RP, Wood PM. Differentiation of axon-related Schwann cells in vitro ascorbic acid regulates basal lamina assembly and myelin formation. Journal of Cell Biology 1987;105(2):1023–34. [4] Plant GW, Currier PF, Cuervo EP, Bates ML, Pressman Y, Bunge MB, et al. Purified adult ensheathing glia fail to myelinate axons under culture conditions that enable Schwann cells to form myelin. Journal of Neuroscience 2002;22(14):6083–91. [5] Visioli F, Reilly MM, Rimoldi M, Solari A, Pareysonc D, CMT-TRIAAL & CMTTRAUK Groups. Vitamin C and Charcot–Marie–Tooth 1A: pharmacokinetic considerations. PharmaNutrition 2013;1:10–2. [6] Levine M, Conry-Cantilena C, Wang Y, Welch RW, Washko PW, Dhariwal KR, et al. Vitamin C pharmacokinetics in healthy volunteers: evidence for a recommended dietary allowance. Proceedings of the National Academy of Sciences of the United States of America 1996;93:3704–9. [7] Micallef J, Attarian S, Dubourg O, Gonnaud PM, Hogrel JY, Stojkovic T, et al. Effect of ascorbic acid in patients with Charcot–Marie–Tooth disease type 1A: a multicentre, randomised, double-blind, placebo-controlled trial. Lancet Neurology 2009;8:1103–10. [8] Pareyson D, Reilly MM, Schenone A, Fabrizi GM, Cavallaro T, Santoro L, et al. Ascorbic acid in Charcot–Marie–Tooth disease type 1A (CMT-TRIAAL and CMTTRAUK): a double-blind randomised trial. Lancet Neurology 2011;10:320–8. [9] Tsukaguchi H, Tokui T, Mackenzie B, Berger UV, Chen XZ, Wang Y, et al. A family of mammalian Na+-dependent L-ascorbic acid transporters. Nature 1999;399:70–5. [10] Daruwala R, Song J, Koh WS, Rumsey SC, Levine M. Cloning and functional characterization of the human sodium-dependent AA transporters hSVCT1 and hSVCT2. FEBS Letters 1999;460(3):480–4. [11] Eck P, Erichsen HC, Taylor JG, Yeager M, Hughes AL, Levine M, et al. Comparison of the genomic structure and variation in the two human sodium-dependent vitamin C transporters, SLC23A1 and SLC23A2. Human Genetics 2004;115: 285–94. [12] Corpe CP, Tu H, Eck P, Wang J, Faulhaber-Walter R, Schnermann J, et al. Vitamin C transporter Slc23a1 links renal reabsorption, vitamin C tissue accumulation, and perinatal survival in mice. Journal of Clinical Investigation 2010;120(4):1069–83.
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