ltal. J. Neurol. Sci. 13.'401 407, 1992
Supplementation of polyunsaturated fatty acids in multiple sclerosis Gallai V., Sarchielli P., Trequattrini A., Murasecco D. Istituto di Clinica delle Malattie Nervose e Mentali, UniversitgJ di Perugia
For several years polyunsaturated fatty acids (PUFAs) and in particular essential fatty acids (EFAs) have been proposed for the treatment of multiple sclerosis (MS). There are contrasting data in literature regarding the effects of the n-6 and the n-3 PUFA series on different aspects of the disease, in particular on the frequency and severity of relapses and platelet function. This can be ascribed to the different criteria of patient selection in relation to the form and severity of disease at the beginning of the various studies. Till now authors have tended to consider the effect of PUFA supplementation only on some clinical aspects of the disease. Modification in the more sensitive indices (immunological or biochemical) of the disease activity should be taken into account and also the influence of dietary lipid intake by patients in relation to n-3 or n-6 EFA supplementation.
Key Words: PUFAs - - Supplementation - - multiple sclerosis
The importance of n-3 and n-6 fatty acids for preventing numerous pathologies, particularly arteriosclerosis, coronary heart disease and hypertension has become evident over the last few years. More recently their therapeutic use has been suggested in some dermatological and neurological pathologies. Linoleic (18:2, n-6) and linolenic (18:3, n-3) acids are respectively precursors of the two series. They have a double bond at the level of C6 or C3 at the methyl end of the molecule. They are essential because there are no specific enzymes able to insert double bonds in these positions. It is also possible to further lengthen the chain at the carboxyl group and to introduce more double bounds in the parent essential fatty acids at this point as well. This process yields long chain fatty acids (20-22 carbon atoms) with 3, 4, 5, 6 double bonds. The two families (n-3 and n-6) formed are fundamental as membrane components and as prostaglandin precursors. Arachidonic acid (5, 8, 11, 14 eicosatetraenoic Received 5 June 1991 - Accepted 30 June 1991
acid) and gamma-linolenic acid (4, 7, 10, 13, 16, 19 docosaesaenoic acid) are the most important derivatives of linolenic acid whereas 5, 8, 11, 14, 17 eicosapentenoic and 4, 7, 10, 13, 16, 19 docosaesaenoic acids are the principal derivatives of alpha-linolenic acid [25]. Arachidonic acid and other EFAs are metabolized mainly by cyclooxygenase and lipooxygenase pathways. Different PG series are the products of cycloxygenase pathways depending on the fatty acid acting as substrate. Prostaglandins of series 2 are formed from linoleic acid and other EFAs of the n-6 series. Prostaglandins of series 3 derive from alpha-linolenic acid and other EFAs of the n-3 series (Table I). Induction of one PG class production inhibits the formation of the other [22]. However, alpha-linolenic acid is an aspecific substrate of cycloxygenase. Indeed, if it is present in large amounts, it should be considered a competitive inhibitor of this enzyme and in this case it is mainly the lipooxygenase pathway that is stimulated [23 ]. 401
The ItalianJounlal of NeurologicalSciences
TABLE 1. Precursors of PGs 0]2 and 3 series. LINOLEIC ACID 18:2 n6 18:3 n6
20:3 n6 ~
PGE1, PGFla
20:4 n6 ~
PGG2
i ! 20:5 n6 ~
PGE2, PGF2a, TXA2 PGI2, PGD2 22:5 n6
LINOLENIC ACID 18:3 n3 18:4 n3
20:4 n3 20:5 n3 - -
22:5 n3 22:6 n3
PGE3, PGF3a, TXA3
EFAs have been proposed for the treatment of MS because their immunosuppressor effects. These effects are achieved not only by the formation of different prostaglandin series but they are also linked to the modification of membrane fluidity induced by these fatty acids which influence receptor binding, the signal transduction pathway and membrane enzyme activity. Prostaglandin
series 2 and 3 have different effects both on blood viscosity and on immune functions. This could have different implications at therapeutic level also in the treatment of MS. The increased incorporation of linoleic acid into the membrane after supplementation induces an aspecific lymphocyte activation that might compete with antigenic activation [24]. Other aspects of immune function (cell-mediated, humoral) are negatively influenced, either in vivo or in vitro, by high levels of EFAs [33,19]. However, this immunosuppressor effect can also be ascribed to inhibition of production of some lymphokines with immunoenhancing action [33]. The effect of EFAs, in particular of the n-3 series, on immune function are reported in Tables II-IV [32,9,15,47,11,31,14,17, 8,7,48,26,39,27,35,49,29,50,41,46,21,37,30,13]. The important impact of high EFA levels on immunocompetence could explain the increasing interest in its use as an alternative therapeutic tool in MS, disease characterized by an immunological disregulation. The interest in EFA supplementation in MS developed from the observation of an association between EFAs intake and MS that was proposed for the first time by Swank [42]. He and others emphasized a different incidence of MS in some geographic areas in relation to dietary EFA intake [43,1,40]. Swank et al. [43] pointed out that diets with a high saturated fatty acids content and a low content of natural antioxidants are involved in the onset of this pathology. Linolenic acid and its polyenoic derivatives are contained in a relatively high proportion in myelin cerebrosides and phospolipids. They are of
TABLE II. Influence of arachidonic acid and linoleic acid supplementation on some immune.fimctions
of experimental animals. Immune function
Animals
Effects
Lymphocyte blastization to PHA, Con A
rats mice mice
reduction
Lymphocyte blastization to alloantigens Cell-mediated citolisis Delayed type hypersensitivity Ability to reject allograft B cell number Serum immunoglobulins Antibody response to thymic-dependent antigens (particularly after primary immunization) IgM, IgG plaque forming cells P G 2 production particularly by monocytes and macrophages NK activity
402
References
(32,9)
mice mice mice rats rats guinea pigs
decrease at high doses, increase at minimal decrease decrease reduction reduction reduction reduction
(15)
rats mice
reduction increase
(31) (7)
mice
reduction
(14)
(47) (11 ) (47,11) (31) (14) (17,8)
Gallai V.: Polyunsaturated fatty acids in M.S.
TABLE Ill. In vitro effect o f E r A on some immune ./'unctions. Immune function
Species
Effects
Lymphocyte response to T dependent mice mitogens (PHA, Con A) guinea pigs human Macrophage phagocytosis mice Surface receptor capping mice Intracetlulartransport and secretion of IgG mouse by plasma-cells (plasmacytoma) (MOPC-31C plasmacytoma cells) B cell response to lipopolisaccarides mice
considerable importance for maintaining optimal stability and integrity of the myelin membranes. Clausen and Hansen [10] showed there was a lower content of linoleic acid in various lipid fractions from different brain areas of MS patients. This could be explained, according to the authors, by a greater intake of cow's milk in the early period of life and in childhood since the E r A content of cow's milk is 5 times lower than that of breast milk. Successive studies on rats have shown the considerable importance of E r A s deficiency during development in regard to the induction of experimental allergic encephalitis. On the other hand, again in rats, supplementing the diet with linoleic acid has a protective effect [18]. It has been argued that an inadequate supply of linoleic acid in the first period of life during the phase of rapid myelin formation could induce lower stability and this could predispose to MS. Nutritional imbalances, regarding particularly lipids and lipid fractions, might also determine an alteration in myelin composition, which could become more sensitive to challenges by pathogens [29]. Reduced E r A membrane content could favor the unmasking of antigenic determinants, for example following a viral infection (paramyxovirus, retrovirus), though to be etiological or causative factors of MS. Since 1969 biochemical studies have shown lower linoleate levels in serum (particularly during relapses) and in CSF and a decrease in the linoleate percentage in membrane phospholipids of platelets, erythrocytes and lymphocytes of MS patients compared to controls [2,3,20,38], though MS patients absorb linoleate like "normal" subjects. A defect in a post-absorptive metabolic pathway has also been suggested. According to the authors, this metabolic defect can be corrected only by supplementing with high amounts of linoleate [6]. Mickel suggested that the reduced C18:2
suppression or enhancement depending from doses and type of veicula (ethanol or albumin) decrease inhibition
References
(26)
(39) (27) (35) (9,49,29)
concentration in body fluids and cellular membranes could be the result of depletion of this fatty acid due to peroxidation processes. The peroxidated E r A s could cause platelet aggregation (as demonstrated in vitro) in CNS post-capillary venules or induce protein denaturation. The denatured proteins could become antigenic and autoimmune processes which might be responsible for disease progression, if not the onset of the pathology [24]. Two research trends have been followed to study the relationship between E r A s and MS. The first designed to assess diet E r A content in relation to severity of the disease and the second to evidence the effect of dietary E r A supplementation on the course of the disease. Swank (1970) reported the results of treatment of MS patients for 20 years with a diet characterized by a reduced intake of saturated fatty acids and a supplement of vegetable oil. Although this study was uncontrolled, he reported that in patients who consumed a smaller quantity of saturated fatty acids and a larger quantity of vegetable oils the clinical worsening'of the condition was reduced [44]. 20 years after, Swank, examining the lipid intake of another 156 MS patients under study from 1949 till 1984, observed that a daily lipid intake of less than 20 g corresponded to a lower mortality (31%) and less worsening of disability. A greater daily lipid intake corresponded to a greater mortality (79%) and a greater worsening of the disability. However early treatment (before the onset of severe disability) produced an improvement in prognosis, particularly in women [45]. According to Fitzgerald et al. the modification of food habits, in particular of E r A intake, and also, in terms of nutrients, of the polyunsaturated/saturated (PUFA/SAFA) ratio in diet, induced a statistically significant increase in the EDSS score of 87 MS patients [16]. The first attempt at E r A supplementation was 403
The Italian Journal of Neurological Sciences
TABLE IV. Effects of n-3 polyunsaturated fato' acids on immune functions. Immune function
Fatty acid administration
Species
Effects
LTB4, LTC4, PGE2 and 6 keto-F1 cx production by stimulated macrophages PGE2 synthesis in spleen and thymus after immune challenge PGE2, LT4 levels and inflammatory essudate following antigenic challenge Delayed type skin reaction 5 lipoxygenase activity of leucocytes Chemotaxis by monocytes and PMN NK activity
fish oil supplementation
mice
reduction
(29)
linolenicacid supplementation
mice
reduction
(50)
EPA and DHA supplementation
mice
reduction
(41)
EPA and DHA supplementation fish oil supplementation fish oil supplementation injection of n3 PUFA triglicerides fish oil supplementation
mice
reduction
(46)
mice
reduction
(41)
man
reduction
(46)
man
decrease
(21)
mice
decrease
(37)
infusion of an emutsion of 10% fish oil fish oil
guinea pigs mice
improvement
(30)
increase
(13)
fish oil
mice
reduce
(13)
Severity of glomerular nephritis in inbred strains with a autoimmune disease similar to LES Survival to injection of an endotoxin Synthesis of IL-1 by peritoneal macrophages INF, cachectin production by macrophages and Kupfer cells
made by Zilkha in 1962. He noticed a clinical improvement in 90 patients with MS after administration of sunflower oil. As this study was unc o n t r o l l e d , M i l l a r et al. of the s a m e r e s e a r c h group, decided to perform a double blind trial, supplementing with linoleic acid another group of 87 MS patients who were not in relapse and with a disability score between 0 and 6 (according the Kurtzke Disability Scale) to confirm these results [341. These patients were r a n d o m l y assigned to two groups: the first treated with linoleic acid, the latter with oleic acid as a placebo. After 28 months no statistically significant differences were observed in the disability scale or on the relapse rate between the groups treated with linoleic acid and with placebo. In contrast the difference was evident regarding 404
References
severity and frequency of clinical relapses, which were greater for the patients who took oleic acid. In 1978 another double blind controlled trial on a group of 116 patients with remitting-relapsing MS for a period of 48 months was conducted by Bates et al. [4]. The patients were assigned randomly to 4 groups: two received linoleic acid, the first as a spread and the latter in capsules with a lower content ( N a u d i c e l l e ) . The third and the fourth groups took oleic acid as a spread or in capsules, respectively. In this second trial the clinical worsening rate was likewise not significantly different among treated and control groups, nor was the frequency of relapses. These relapses were shorter only in the patients who received higher doses of linoleic acid. In a third double blind trial Paty et al. [36] studied the effects of linoleic acid supplementation
Gallai V.: Polyunsaturated fatty acids in M.S.
in 76 MS patients for a period of 30 months. The supplementation had no influence on the neurological course, relapse rate or severity. Dworkin et al. [12] reviewed the results of the 3 trials reported above. Because the doses and procedures followed in these 3 studies were similar a single sample of 181 patients (90 who took linoleic acid and 91 who took oleic acid as a placebo) was obtained from these 3 trials. Statistical analysis showed that, in terms of EDSS modification, the patients with little or no disability at the beginning of the study and who had been treated with linoleic acid had less increase of disability and a lower relapse rate than the control group irrespective of the disability rating. There were no statistically significant differences between the two groups in regard to the number of relapses per year. The reproducibility of linoleic acid effects on relapse severity and duration was also greater in patients with a lower disability score and a shorter disease duration. Only one study has been conducted on the effects of EFA of n-3 series on the course of MS. In 1988 Bates et al. [5] conducted a double blind controlled trial on 312 patients supplementing with Maxepa, a fish oil containing 18% of docosaesaenoic acid and 12% of eicosapentaenoic acid. The results showed no differences between the two groups treated with Maxepa and a placebo respectively, but a positive trend for the group treated with n-3 PUFA for all the characteristics considered (relapse duration, frequency and severity). Evidence of a clinical improvement or at least of no clinical worsening emerged. In a recent study on the effects of EFAs on platelet function in the course of MS Gregor et al. [28] assessed the effects of supplementing for 6 weeks with Efamol oil which has a high content of linoleic (70%) and gamma-linoleic (9%) acids, on the function and glycoprotein composition of platelets in MS patients. They found no modification of platelet aggregability after ADP and collagen stimulation and of 125I-fibrinogen binding. Platelet aggregability was increased in patients compared to the controls and there was a statistically significant difference only following thrombin stimulation after Efamol treatment. These findings contrast with those obtained in experimental animals (rats) where supplementing the diet with linoleic acid induced a reduction of platelet aggregability, modification of the fatty acid composition of membrane phospholipids and prostaglandin synthesis. This was accompanied by an increased binding of fibrinogen which could be correlated to the increased membrane fluidity due to a greater content of linoleic acid in platelet membrane in the supplemented animals. No studies have been done on the modification of
immune function following EFA supplementation, nor have PG levels in urine, serum or cellular membrane been determined. All these aspects should be the object of further investigations.
Conclusions The studies on the therapeutic use of PUFAs in MS to date have not always evidenced positive effects on the course of disease. This can be attributed mostly to the different criteria of patient selection (some with remitting form, others with progressive form) and to the different disability rating of patients at the beginning of the study. The studies were not long enough to evidence real modifications in the clinical course. Perhaps variations in biochemical or immunological indicators of the disease activity might show earlier modifications of the course. The n-3 and n-6 PUFA series could have different effects at membrane level, in regard to platelet activity and immune function, because they are precursors of PG classes of different and sometimes antagonistic activity. In spite of the incomplete agreement on the results of various studies and the need for further double blind controlled trials (chiefly for the n-3 PUFAs and EFAs) a modification of the food habits, in particular the EFAs and PUFAs/SAFAs ratio, and a periodic supplementing with the EFAs of both series could still support immunosuppressive therapy which should certainly not be considered a long term therapy. This could be more effective, especially in those patients without severe disability, for reducing relapse frequency and severity. Further clinical and experimental studies are needed to assess the influence of n-3 and n-6 PUFAs, and in particular EFAs, or of both series together, on different aspects of the disease (immune function, platelet activity, clinical course). Efforts should be made to identify the optimal ratio between the two classes with the greatest therapeutic effect. Finally the importance of lipid intake assessment (particularly EFAs and the PUFA/SAFA ratio) of MS patients should be considered before undertaking supplementation trials. The need to assure a similar intake of EFAs from both series, also by a more adequate diet, emerges from the contrasting data in literature. In fact the lack of a similar dietary lipid intake between the supplemented and the control groups could have an impact, even if minimal, on the immune function and on some aspects of the pathology, irrespective of the supplementation, which could in some way influence the results. 405
The Italian Journal (ff Neurological Sciences
Sommario
Da diversi anni gli acidi grassi poliinsaturi (PUFAs) e, in particolare, gli acidi grassi essenziali (EFAs), sono stati proposti per il trattamento della Sclerosi Multipla (MS). Vi sono dati contrastanti riguardo gli effetti dei P U F A s delle serie n-3 ed n-6 su differenti aspetti della malattia, quali la frequenza e la gravitgt delle riacutizzazioni e la funzione piastrinica. Ci6 puO essere attribuito ai differenti criteri utilizzati dagli Autori per la selezione dei pazienti. Fino a questo momento gli studi si sono limitati a considerare gli effetti della supplementazione di P U F A s solo su alcuni aspetti clinici della MS. Dovrebbero essere invece considerati indicatori piit sensibili dell'attivitgt della malattia, come quelli immunologici e quelli biochimici, valutando gli effetti della supplementazione con E F A s delle serie n-3 o n-6 su questi indicatori, oltrechi sull'andamento clinico della patologia.
Address reprint requests to: Prof. Virgilio Gallai Clinica delle Malattie Nervose e Mentali Casella Postale n. 1427 - Succ. 3 06126 Perugia
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407
Supplementation of polyunsaturated fatty acids in multiple sclerosis Gallai V., Sarchielli P., Trequattrini A., Murasecco D. Istituto di Clinica delle Malattie Nervose e Mentali, UniversitgJ di Perugia
For several years polyunsaturated fatty acids (PUFAs) and in particular essential fatty acids (EFAs) have been proposed for the treatment of multiple sclerosis (MS). There are contrasting data in literature regarding the effects of the n-6 and the n-3 PUFA series on different aspects of the disease, in particular on the frequency and severity of relapses and platelet function. This can be ascribed to the different criteria of patient selection in relation to the form and severity of disease at the beginning of the various studies. Till now authors have tended to consider the effect of PUFA supplementation only on some clinical aspects of the disease. Modification in the more sensitive indices (immunological or biochemical) of the disease activity should be taken into account and also the influence of dietary lipid intake by patients in relation to n-3 or n-6 EFA supplementation.
Key Words: PUFAs - - Supplementation - - multiple sclerosis
The importance of n-3 and n-6 fatty acids for preventing numerous pathologies, particularly arteriosclerosis, coronary heart disease and hypertension has become evident over the last few years. More recently their therapeutic use has been suggested in some dermatological and neurological pathologies. Linoleic (18:2, n-6) and linolenic (18:3, n-3) acids are respectively precursors of the two series. They have a double bond at the level of C6 or C3 at the methyl end of the molecule. They are essential because there are no specific enzymes able to insert double bonds in these positions. It is also possible to further lengthen the chain at the carboxyl group and to introduce more double bounds in the parent essential fatty acids at this point as well. This process yields long chain fatty acids (20-22 carbon atoms) with 3, 4, 5, 6 double bonds. The two families (n-3 and n-6) formed are fundamental as membrane components and as prostaglandin precursors. Arachidonic acid (5, 8, 11, 14 eicosatetraenoic Received 5 June 1991 - Accepted 30 June 1991
acid) and gamma-linolenic acid (4, 7, 10, 13, 16, 19 docosaesaenoic acid) are the most important derivatives of linolenic acid whereas 5, 8, 11, 14, 17 eicosapentenoic and 4, 7, 10, 13, 16, 19 docosaesaenoic acids are the principal derivatives of alpha-linolenic acid [25]. Arachidonic acid and other EFAs are metabolized mainly by cyclooxygenase and lipooxygenase pathways. Different PG series are the products of cycloxygenase pathways depending on the fatty acid acting as substrate. Prostaglandins of series 2 are formed from linoleic acid and other EFAs of the n-6 series. Prostaglandins of series 3 derive from alpha-linolenic acid and other EFAs of the n-3 series (Table I). Induction of one PG class production inhibits the formation of the other [22]. However, alpha-linolenic acid is an aspecific substrate of cycloxygenase. Indeed, if it is present in large amounts, it should be considered a competitive inhibitor of this enzyme and in this case it is mainly the lipooxygenase pathway that is stimulated [23 ]. 401
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TABLE 1. Precursors of PGs 0]2 and 3 series. LINOLEIC ACID 18:2 n6 18:3 n6
20:3 n6 ~
PGE1, PGFla
20:4 n6 ~
PGG2
i ! 20:5 n6 ~
PGE2, PGF2a, TXA2 PGI2, PGD2 22:5 n6
LINOLENIC ACID 18:3 n3 18:4 n3
20:4 n3 20:5 n3 - -
22:5 n3 22:6 n3
PGE3, PGF3a, TXA3
EFAs have been proposed for the treatment of MS because their immunosuppressor effects. These effects are achieved not only by the formation of different prostaglandin series but they are also linked to the modification of membrane fluidity induced by these fatty acids which influence receptor binding, the signal transduction pathway and membrane enzyme activity. Prostaglandin
series 2 and 3 have different effects both on blood viscosity and on immune functions. This could have different implications at therapeutic level also in the treatment of MS. The increased incorporation of linoleic acid into the membrane after supplementation induces an aspecific lymphocyte activation that might compete with antigenic activation [24]. Other aspects of immune function (cell-mediated, humoral) are negatively influenced, either in vivo or in vitro, by high levels of EFAs [33,19]. However, this immunosuppressor effect can also be ascribed to inhibition of production of some lymphokines with immunoenhancing action [33]. The effect of EFAs, in particular of the n-3 series, on immune function are reported in Tables II-IV [32,9,15,47,11,31,14,17, 8,7,48,26,39,27,35,49,29,50,41,46,21,37,30,13]. The important impact of high EFA levels on immunocompetence could explain the increasing interest in its use as an alternative therapeutic tool in MS, disease characterized by an immunological disregulation. The interest in EFA supplementation in MS developed from the observation of an association between EFAs intake and MS that was proposed for the first time by Swank [42]. He and others emphasized a different incidence of MS in some geographic areas in relation to dietary EFA intake [43,1,40]. Swank et al. [43] pointed out that diets with a high saturated fatty acids content and a low content of natural antioxidants are involved in the onset of this pathology. Linolenic acid and its polyenoic derivatives are contained in a relatively high proportion in myelin cerebrosides and phospolipids. They are of
TABLE II. Influence of arachidonic acid and linoleic acid supplementation on some immune.fimctions
of experimental animals. Immune function
Animals
Effects
Lymphocyte blastization to PHA, Con A
rats mice mice
reduction
Lymphocyte blastization to alloantigens Cell-mediated citolisis Delayed type hypersensitivity Ability to reject allograft B cell number Serum immunoglobulins Antibody response to thymic-dependent antigens (particularly after primary immunization) IgM, IgG plaque forming cells P G 2 production particularly by monocytes and macrophages NK activity
402
References
(32,9)
mice mice mice rats rats guinea pigs
decrease at high doses, increase at minimal decrease decrease reduction reduction reduction reduction
(15)
rats mice
reduction increase
(31) (7)
mice
reduction
(14)
(47) (11 ) (47,11) (31) (14) (17,8)
Gallai V.: Polyunsaturated fatty acids in M.S.
TABLE Ill. In vitro effect o f E r A on some immune ./'unctions. Immune function
Species
Effects
Lymphocyte response to T dependent mice mitogens (PHA, Con A) guinea pigs human Macrophage phagocytosis mice Surface receptor capping mice Intracetlulartransport and secretion of IgG mouse by plasma-cells (plasmacytoma) (MOPC-31C plasmacytoma cells) B cell response to lipopolisaccarides mice
considerable importance for maintaining optimal stability and integrity of the myelin membranes. Clausen and Hansen [10] showed there was a lower content of linoleic acid in various lipid fractions from different brain areas of MS patients. This could be explained, according to the authors, by a greater intake of cow's milk in the early period of life and in childhood since the E r A content of cow's milk is 5 times lower than that of breast milk. Successive studies on rats have shown the considerable importance of E r A s deficiency during development in regard to the induction of experimental allergic encephalitis. On the other hand, again in rats, supplementing the diet with linoleic acid has a protective effect [18]. It has been argued that an inadequate supply of linoleic acid in the first period of life during the phase of rapid myelin formation could induce lower stability and this could predispose to MS. Nutritional imbalances, regarding particularly lipids and lipid fractions, might also determine an alteration in myelin composition, which could become more sensitive to challenges by pathogens [29]. Reduced E r A membrane content could favor the unmasking of antigenic determinants, for example following a viral infection (paramyxovirus, retrovirus), though to be etiological or causative factors of MS. Since 1969 biochemical studies have shown lower linoleate levels in serum (particularly during relapses) and in CSF and a decrease in the linoleate percentage in membrane phospholipids of platelets, erythrocytes and lymphocytes of MS patients compared to controls [2,3,20,38], though MS patients absorb linoleate like "normal" subjects. A defect in a post-absorptive metabolic pathway has also been suggested. According to the authors, this metabolic defect can be corrected only by supplementing with high amounts of linoleate [6]. Mickel suggested that the reduced C18:2
suppression or enhancement depending from doses and type of veicula (ethanol or albumin) decrease inhibition
References
(26)
(39) (27) (35) (9,49,29)
concentration in body fluids and cellular membranes could be the result of depletion of this fatty acid due to peroxidation processes. The peroxidated E r A s could cause platelet aggregation (as demonstrated in vitro) in CNS post-capillary venules or induce protein denaturation. The denatured proteins could become antigenic and autoimmune processes which might be responsible for disease progression, if not the onset of the pathology [24]. Two research trends have been followed to study the relationship between E r A s and MS. The first designed to assess diet E r A content in relation to severity of the disease and the second to evidence the effect of dietary E r A supplementation on the course of the disease. Swank (1970) reported the results of treatment of MS patients for 20 years with a diet characterized by a reduced intake of saturated fatty acids and a supplement of vegetable oil. Although this study was uncontrolled, he reported that in patients who consumed a smaller quantity of saturated fatty acids and a larger quantity of vegetable oils the clinical worsening'of the condition was reduced [44]. 20 years after, Swank, examining the lipid intake of another 156 MS patients under study from 1949 till 1984, observed that a daily lipid intake of less than 20 g corresponded to a lower mortality (31%) and less worsening of disability. A greater daily lipid intake corresponded to a greater mortality (79%) and a greater worsening of the disability. However early treatment (before the onset of severe disability) produced an improvement in prognosis, particularly in women [45]. According to Fitzgerald et al. the modification of food habits, in particular of E r A intake, and also, in terms of nutrients, of the polyunsaturated/saturated (PUFA/SAFA) ratio in diet, induced a statistically significant increase in the EDSS score of 87 MS patients [16]. The first attempt at E r A supplementation was 403
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TABLE IV. Effects of n-3 polyunsaturated fato' acids on immune functions. Immune function
Fatty acid administration
Species
Effects
LTB4, LTC4, PGE2 and 6 keto-F1 cx production by stimulated macrophages PGE2 synthesis in spleen and thymus after immune challenge PGE2, LT4 levels and inflammatory essudate following antigenic challenge Delayed type skin reaction 5 lipoxygenase activity of leucocytes Chemotaxis by monocytes and PMN NK activity
fish oil supplementation
mice
reduction
(29)
linolenicacid supplementation
mice
reduction
(50)
EPA and DHA supplementation
mice
reduction
(41)
EPA and DHA supplementation fish oil supplementation fish oil supplementation injection of n3 PUFA triglicerides fish oil supplementation
mice
reduction
(46)
mice
reduction
(41)
man
reduction
(46)
man
decrease
(21)
mice
decrease
(37)
infusion of an emutsion of 10% fish oil fish oil
guinea pigs mice
improvement
(30)
increase
(13)
fish oil
mice
reduce
(13)
Severity of glomerular nephritis in inbred strains with a autoimmune disease similar to LES Survival to injection of an endotoxin Synthesis of IL-1 by peritoneal macrophages INF, cachectin production by macrophages and Kupfer cells
made by Zilkha in 1962. He noticed a clinical improvement in 90 patients with MS after administration of sunflower oil. As this study was unc o n t r o l l e d , M i l l a r et al. of the s a m e r e s e a r c h group, decided to perform a double blind trial, supplementing with linoleic acid another group of 87 MS patients who were not in relapse and with a disability score between 0 and 6 (according the Kurtzke Disability Scale) to confirm these results [341. These patients were r a n d o m l y assigned to two groups: the first treated with linoleic acid, the latter with oleic acid as a placebo. After 28 months no statistically significant differences were observed in the disability scale or on the relapse rate between the groups treated with linoleic acid and with placebo. In contrast the difference was evident regarding 404
References
severity and frequency of clinical relapses, which were greater for the patients who took oleic acid. In 1978 another double blind controlled trial on a group of 116 patients with remitting-relapsing MS for a period of 48 months was conducted by Bates et al. [4]. The patients were assigned randomly to 4 groups: two received linoleic acid, the first as a spread and the latter in capsules with a lower content ( N a u d i c e l l e ) . The third and the fourth groups took oleic acid as a spread or in capsules, respectively. In this second trial the clinical worsening rate was likewise not significantly different among treated and control groups, nor was the frequency of relapses. These relapses were shorter only in the patients who received higher doses of linoleic acid. In a third double blind trial Paty et al. [36] studied the effects of linoleic acid supplementation
Gallai V.: Polyunsaturated fatty acids in M.S.
in 76 MS patients for a period of 30 months. The supplementation had no influence on the neurological course, relapse rate or severity. Dworkin et al. [12] reviewed the results of the 3 trials reported above. Because the doses and procedures followed in these 3 studies were similar a single sample of 181 patients (90 who took linoleic acid and 91 who took oleic acid as a placebo) was obtained from these 3 trials. Statistical analysis showed that, in terms of EDSS modification, the patients with little or no disability at the beginning of the study and who had been treated with linoleic acid had less increase of disability and a lower relapse rate than the control group irrespective of the disability rating. There were no statistically significant differences between the two groups in regard to the number of relapses per year. The reproducibility of linoleic acid effects on relapse severity and duration was also greater in patients with a lower disability score and a shorter disease duration. Only one study has been conducted on the effects of EFA of n-3 series on the course of MS. In 1988 Bates et al. [5] conducted a double blind controlled trial on 312 patients supplementing with Maxepa, a fish oil containing 18% of docosaesaenoic acid and 12% of eicosapentaenoic acid. The results showed no differences between the two groups treated with Maxepa and a placebo respectively, but a positive trend for the group treated with n-3 PUFA for all the characteristics considered (relapse duration, frequency and severity). Evidence of a clinical improvement or at least of no clinical worsening emerged. In a recent study on the effects of EFAs on platelet function in the course of MS Gregor et al. [28] assessed the effects of supplementing for 6 weeks with Efamol oil which has a high content of linoleic (70%) and gamma-linoleic (9%) acids, on the function and glycoprotein composition of platelets in MS patients. They found no modification of platelet aggregability after ADP and collagen stimulation and of 125I-fibrinogen binding. Platelet aggregability was increased in patients compared to the controls and there was a statistically significant difference only following thrombin stimulation after Efamol treatment. These findings contrast with those obtained in experimental animals (rats) where supplementing the diet with linoleic acid induced a reduction of platelet aggregability, modification of the fatty acid composition of membrane phospholipids and prostaglandin synthesis. This was accompanied by an increased binding of fibrinogen which could be correlated to the increased membrane fluidity due to a greater content of linoleic acid in platelet membrane in the supplemented animals. No studies have been done on the modification of
immune function following EFA supplementation, nor have PG levels in urine, serum or cellular membrane been determined. All these aspects should be the object of further investigations.
Conclusions The studies on the therapeutic use of PUFAs in MS to date have not always evidenced positive effects on the course of disease. This can be attributed mostly to the different criteria of patient selection (some with remitting form, others with progressive form) and to the different disability rating of patients at the beginning of the study. The studies were not long enough to evidence real modifications in the clinical course. Perhaps variations in biochemical or immunological indicators of the disease activity might show earlier modifications of the course. The n-3 and n-6 PUFA series could have different effects at membrane level, in regard to platelet activity and immune function, because they are precursors of PG classes of different and sometimes antagonistic activity. In spite of the incomplete agreement on the results of various studies and the need for further double blind controlled trials (chiefly for the n-3 PUFAs and EFAs) a modification of the food habits, in particular the EFAs and PUFAs/SAFAs ratio, and a periodic supplementing with the EFAs of both series could still support immunosuppressive therapy which should certainly not be considered a long term therapy. This could be more effective, especially in those patients without severe disability, for reducing relapse frequency and severity. Further clinical and experimental studies are needed to assess the influence of n-3 and n-6 PUFAs, and in particular EFAs, or of both series together, on different aspects of the disease (immune function, platelet activity, clinical course). Efforts should be made to identify the optimal ratio between the two classes with the greatest therapeutic effect. Finally the importance of lipid intake assessment (particularly EFAs and the PUFA/SAFA ratio) of MS patients should be considered before undertaking supplementation trials. The need to assure a similar intake of EFAs from both series, also by a more adequate diet, emerges from the contrasting data in literature. In fact the lack of a similar dietary lipid intake between the supplemented and the control groups could have an impact, even if minimal, on the immune function and on some aspects of the pathology, irrespective of the supplementation, which could in some way influence the results. 405
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Sommario
Da diversi anni gli acidi grassi poliinsaturi (PUFAs) e, in particolare, gli acidi grassi essenziali (EFAs), sono stati proposti per il trattamento della Sclerosi Multipla (MS). Vi sono dati contrastanti riguardo gli effetti dei P U F A s delle serie n-3 ed n-6 su differenti aspetti della malattia, quali la frequenza e la gravitgt delle riacutizzazioni e la funzione piastrinica. Ci6 puO essere attribuito ai differenti criteri utilizzati dagli Autori per la selezione dei pazienti. Fino a questo momento gli studi si sono limitati a considerare gli effetti della supplementazione di P U F A s solo su alcuni aspetti clinici della MS. Dovrebbero essere invece considerati indicatori piit sensibili dell'attivitgt della malattia, come quelli immunologici e quelli biochimici, valutando gli effetti della supplementazione con E F A s delle serie n-3 o n-6 su questi indicatori, oltrechi sull'andamento clinico della patologia.
Address reprint requests to: Prof. Virgilio Gallai Clinica delle Malattie Nervose e Mentali Casella Postale n. 1427 - Succ. 3 06126 Perugia
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