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Plasma homocysteine levels in young male patients in the exacerbation and remission phase of schizophrenia
Progress in Neuro-Psychopharmacology & Biological Psychiatry 32 (2008) 1921–1926
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Progress in Neuro-Psychopharmacology & Biological Psychiatry j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / p n p b p
Plasma homocysteine levels in young male patients in the exacerbation and remission phase of schizophrenia Nataša D. Petronijević a,⁎, Nevena V. Radonjić a, Maja D. Ivković b, Dragan Marinković b, Vesna D. Piperski c, Bogdan M. Đuričić a, Vladimir R. Paunović b a b c
Institute of Medical and Clinical Biochemistry, School of Medicine, Pasterova 2, 11000 Belgrade, Serbia Institute of Psychiatry, School of Medicine, Pasterova 2, 11000 Belgrade, Serbia Present address: Medical Academy US Medical School, Kumodraška 261, 11000 Beograd, Serbia
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Article history: Received 17 March 2008 Received in revised form 10 September 2008 Accepted 11 September 2008 Available online 15 September 2008 Keywords: B12 Folate Homocysteine Negative symptoms Positive symptoms Schizophrenia
a b s t r a c t High levels of homocysteine (Hcy) were suggested to contribute to the pathogenesis of schizophrenia. Recent investigations have shown that treatment with folic acid, vitamin B-12 and pyridoxine are effective in reducing Hcy levels while concomitantly reducing the score of positive and negative symptoms in schizophrenic patients. In addition to the availability of nutrients (mainly folate, vitamins B6 and B12), plasma Hcy concentrations are dependent on complex metabolic regulation that could be disrupted in schizophrenia. This study was designed to test the influence of disease activity on plasma Hcy levels. Plasma Hcy concentrations were measured in male chronic schizophrenic patients with a predominantly positive (SCH (+)) or predominantly negative (SCH (−)) syndrome in schizophrenia immediately upon admission to the hospital (exacerbation phase) and one month later (remission phase). During this period patients received antipsychotic medications without vitamin therapy. The effects of age, duration of illness, folate and B12 concentrations, as well as smoking and coffee consumption habits on the observed changes were evaluated. Age- and sex-matched subjects were included in the control group. In the control group plasma Hcy concentration was 8.75 ± 1.84 μmol/L. In the exacerbation phase plasma Hcy concentrations were significantly increased both in SCH (+) (14.91 ± 6.19 μmol/L) and SCH (−) groups (12.8 ± 3.27 μmol/L). There was no difference in plasma Hcy concentrations between SCH (+) and SCH (−) patients. Serum folate and B12 concentrations were not significantly different in any of the investigated groups of subjects. The plasma Hcy concentrations could not be correlated with age, duration of illness, the score of positive symptoms or the concentration of folate and vitamin B12. A positive correlation was found between plasma Hcy level and score of negative symptoms in both groups of patients. No correlation was found between smoking or coffee consumption habits and plasma Hcy concentrations. All patients exhibited decreased plasma Hcy levels in the remission phase of the illness, with a mean decrease of 2.68 ± 1.57 μmol/L. Folate and B12 levels did not differ in the exacerbation and remission phases of the illness. The significant decrease of plasma Hcy levels, without changes in folate and vitamin B12 concentrations in the remission phase of schizophrenia, could indicate an influence of a pathogenetic process involved in schizophrenia on Hcy metabolism. © 2008 Elsevier Inc. All rights reserved.
1. Introduction Homocysteine (Hcy) is a sulfur containing amino acid formed during the metabolism of the essential amino acid methionine. In most tissues, Hcy may be remethylated to methionine by the enzyme
Abbreviations: BMI, body mass index; EDTA, ethylenediaminetetraacetic acid; EIA, enzyme immunoassay; Hcy, homocysteine; HPA, hypothalamic-pituitary-adrenal; MTHFR, methylenetetrahydrofolate reductase; PANSS, positive and negative syndrome scale; PTSD, posttraumatic stress disorder; SCH, schizophrenia. ⁎ Corresponding author. Tel./fax: +381 11 2682 953. E-mail address: [email protected] (N.D. Petronijević). 0278-5846/$ – see front matter © 2008 Elsevier Inc. All rights reserved. doi:10.1016/j.pnpbp.2008.09.009
methionine synthase by a reaction where vitamin B12 is a cofactor and methyltetrahydrofolate is a substrate. Elevated level of plasma total Hcy was found to be a risk factor for cardiovascular (Nygård et al., 1995) and cerebrovascular disease (Furie and Kelly, 2006). Increased total plasma Hcy levels have also been reported in several neuropsychiatric disorders including Alzheimer's disease (Seshadri et al., 2002), Parkinson's disease (O'Suilleabhain et al., 2006; Hassin-Baer et al., 2006; Todorović et al., 2006), depression (Bottiglieri et al., 2000; Folstein et al., 2007), bipolar disorder (Osher et al., 2004), schizophrenia and schizophrenia-like psychosis (Regland et al., 1995; Susser et al., 1998; Levine et al., 2002). Markedly elevated plasma Hcy levels have been found in young male schizophrenic patients (Levine et al., 2002), a finding that has
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been replicated in a group of newly admitted schizophrenic patients (Applebaum et al., 2004), as well as in adolescents (age 14–21 years) suffering from schizophrenia (Adler Nevo et al., 2006). On the other hand, reports regarding hyperhomocysteinemia in female schizophrenic patients have not been consistent. Haidemenos et al. (2007) did not find gender and age differences of plasma Hcy levels in schizophrenic patients, suggesting that hyperhomocystinemia may be a risk factor for schizophrenia regardless of gender. However, another study failed to detect elevated Hcy levels in young schizophrenic women (Reif et al., 2003). Notably, elevated Hcy levels among pregnant women have been found to increase the risk of their children developing schizophrenia (Brown et al., 2007). High levels of Hcy have been suggested to contribute to the pathogenesis of schizophrenia (Levine et al., 2005). Recently, studies have suggested that reducing Hcy levels is an effective strategy to ameliorate clinical symptoms in schizophrenic patients. Levine et al. (2006) has shown that treatment with folic acid, vitamin B-12 and pyridoxine is effective in reducing Hcy levels as well as the scores of positive and negative symptoms in schizophrenic patients with starting plasma Hcy concentrations above 15 μmol/L. The results of a preliminary study by Miodownik et al. (2007) indicated the effectiveness of high dose vitamin B6 in decreasing serum Hcy levels in male schizophrenic patients. The rationale for the current study was to determine whether the elevated homocysteine level reported in this disorder is a causative factor or is a result of a pathophysiologic process of schizophrenia and if such an elevated level is a state or a trait marker for this disease. Plasma Hcy levels were measured in young male chronic schizophrenic patients with predominantly positive or predominantly negative symptoms, during the exacerbation phase of the illness and one month later, in the remission phase. During this period patients received neuroleptic medication without vitamin supplements. The use of different antipsychotics showed no trend for association with increased Hcy (Levine et al., 2005). The effects of age, duration of illness, folate and vitamin B12 concentrations, as well as smoking and coffee consumption habits, on observed changes were evaluated. 2. Patients and methods Fifty-two male patients from the Institute of Psychiatry, School of Medicine, University of Belgrade, which met DSM-IV criteria (American Psychiatric Association, 1994) for schizophrenia and 20 healthy control male healthy volunteers recruited from the staff of the Institute of Psychiatry and Institute of Medical and Clinical Biochemistry were included in the study. Three of the recruited patients did not agree to participate in the study and five others were not in the remission phase after one month. The final patient group consisted of 44 subjects. In the SCH group, psychopathology was assessed by Positive and Negative Syndrome Scale (PANSS). All patients were divided into two groups: with either a predominantly positive [SCH (+)] or a predominantly negative [SCH (−)] syndrome in schizophrenia (SCH) (Kay et al., 1987). Empirically established percentile ranks derived from raw and standardized T-scores on the PANSS profile form were used to suit research objectives. This procedure declares that a person with decidedly high positive or negative profile scores is at the extremes of the schizophrenic sample-distribution curve for the PANSS composite scale. The authors used the most rigorous criterion. Consequently, only those patients ranked above the 95th percentile (composite score of 11 and higher) were considered SCH (+), whereas those ranking below the 25th percentile (composite scores of −15 and lower) were considered SCH (−). All subjects were otherwise medically healthy and no subjects had histories of cardiovascular or neurological disease, diabetes, or a history of substance abuse or dependence. Written informed consent was obtained from each subject after a complete description of the study was provided. Patients were introduced into the study in the exacerbation phase. Twenty-six SCH patients had a predominantly positive (21–
50 years old; mean 35.28 ± 8.28) and eighteen a predominantly negative (21–50 years old; mean 33.78 ± 9.37) syndrome. All patients included in the study were treated with antipsychotic drugs (mean neuroleptic dose in chlorpromazine equivalents was 276 ± 161 mg per day; range: 100–630 mg per day). Patients were also grouped as smokers or non-smokers (smokers consumed 16.2 ± 3.5 cigarettes per day) and according to their coffee consumption habits (coffee consumers drink at least one cup of coffee per day). The blood samples were collected immediately upon admission to the hospital in the exacerbation phase and again one month later in the remission phase of illness. All biochemical measurements were conducted blind to diagnostic status. During the study all patients were on regular hospital diet, and there were no clinically relevant symptoms of malnutrition. Control subjects (n = 20) were male healthy volunteers (25– 45 years old; mean 33 ± 8.1). Because non-psychotic, first-degree relatives of SCH probands may carry a genetic diathesis to SCH (Amin et al., 1999), the subjects included in the control group were required to be free of any major psychiatric disorder in their first-degree relatives. Additionally, they were required to have a negative family history of dementia, mental retardation, and Huntington's disease in first-degree relatives. Smoking was prohibited after 11 p.m. before the next day's blood draw. None of the subjects had taken any vitamin supplements three months prior to being included in the study. Subjects also did not take any medications (carbamazepine, phenytoin, anticonvulsants or 6-azauridine triacetate) that are known to cause an elevation of plasma Hcy levels. Food consumption can affect circulating Hcy levels and protein rich meals were avoided late in the day before blood sampling. 2.1. Collection of blood samples and biochemical analyses Blood samples were taken from antecubital vein between 7 and 8 a.m. into a vacutainer containing 1 mL 1% EDTA as an anticoagulant. The samples were kept on ice (maximum of 1 h) and plasma and blood cells were separated by centrifugation (15 min, 3000 rpm). Plasma samples were stored at −80 °C until Hcy measurements. Hcy levels were measured by Bio-Rad Microplate Enzyme Immunoassay Homocysteine Test (EIA) (Bio-Rad Laboratories, Inc., Hercules, CA, USA). Serum levels of folate and vitamin B12 were determined by competitive chemiluminescent assays with an IMMULITE 1000 analyzer (Siemens Healthcare Diagnostics, NY, USA). The data sets were tested for normal distribution by the Kolmogorov– Smirnov test. All data sets were within normal distribution and analyzed using the analysis of variance (ANOVA) followed by Fisher's test for multiple comparison. Associations between age, duration of illness, clinical rating scale scores, serum folate and B12 and levels of plasma Hcy were tested by Pearson's correlation coefficient. For assessment of
Fig. 1. Plasma Hcy levels in control subjects (n = 20) and patients suffering from schizophrenia included in SCH (+) (n = 26) and SCH (−) (n = 18) group. ⁎p b 0.01 in SCH (+) group vs. controls and p b 0.05 in SCH (−) group vs. controls.
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Fig. 3. Plasma Hcy concentration in all SCH patients in the exacerbation phase upon admission and in the remission phase of illness one month later.
Fig. 2. Correlation of plasma Hcy levels with the score of negative (A and B) symptoms in SCH (+) (n = 26) and SCH (−) (n = 18) patients.
smoking or coffee consumption influence on the plasma Hcy levels in patients with predominantly positive or negative syndrome in SCH, the Student's t-test was used. Significance of plasma Hcy and serum folate and vitamin B12 concentration changes in exacerbation and remission phase of illness was tested with paired t-test. Results are presented as mean ±SD and pb 0.05 was regarded as significant for all comparisons. 3. Results When compared to healthy controls, plasma Hcy concentrations (Fig. 1) were significantly elevated in both SCH (+) and SCH (−) patients immediately upon admission to the hospital (df = 2; p = 0.0001). Although SCH (+) patients had higher plasma Hcy concentrations than SCH (−) patients, the difference was not statistically significant. Statistically significant correlations between plasma Hcy concentrations and age (R = 0.17, p = 0.65 for SCH (+); R = 0.48, p = 0.19 for SCH (−)) or duration of illness (R = 0.37, p = 0.24 for SCH (+); R = 0.51, p = 0.25 for SCH (−)) were not found in either SCH (+) or SCH (−) patients. Correlations of plasma Hcy levels and the scores of negative symptoms are presented on Fig. 2. A significant correlation was found
Table 1 Serum folate and vitamin B12 levels in control group and SCH (+) and SCH (−) groups of patients immediately upon admission to the hospital Controls
SCH (+)
SCH (−)
n = 20
n = 26
n = 18
SCH (+) vs. SCH (−) vs. SCH (+) vs. control control SCH (−)
Folate (ng/mL) 8.19 ± 1.69 6.35 ± 1.69 7.67 ± 1.89 p = 0.08 B12 (pg/mL) 397 ± 241 516 ± 296 363 ± 163 p = 0.45
p = 0.72 p = 0.61
Values are represented as mean ± SD for specified number of subjects.
p = 0.21 p = 0.32
between plasma Hcy levels and scores of negative symptoms in SCH (+) as well as in SCH (−) group (Fig. 2). At the same time there was no significant correlation between plasma Hcy levels and scores of positive symptoms in both SCH (+) (R = 0.28, p = 0.166) and SCH (−) (R = −0.195, p = 0.43) groups. The concentrations of serum folate and vitamin B12 did not differ between investigated groups of subjects (Table 1). Although the concentration of folate was lower in the SCH (+) group compared to the control group, the difference did not reach statistical significance. There was no significant correlation between plasma Hcy and folate (R = − 0.317, df = 43, p = 0.14) or vitamin B12 (R = − 0.314, df = 43, p = 0.154) levels in patients upon admission to the hospital. Comparison of plasma Hcy levels measured immediately upon admission to the hospital and one month later is presented at Fig. 3. The mean change of plasma Hcy level between those two measurements was 2.68 ± 1.57 μmol/L, and the difference was statistically significant (p = 0.00478: t = 10.42; df = 43). Regardless of the starting Hcy level, decreased plasma Hcy levels were observed in every patient tested. In the remission phase mean plasma Hcy level in SCH (+) group was 11.18 ± 4.7 μmol/L (p = 0.14 comparing to control group) and in SCH (−) group 10.07 ± 2.51 μmol/L (p = 0.21 comparing to control group), suggesting that after one month of treatment, regardless of positive or negative symptomatology upon admission, patient plasma Hcy levels were indistinguishable from those of the control group. The levels of folate and vitamin B12 did not differ between patients immediately after admission to the hospital. As investigated by paired t-test, there were no significant differences in the between levels of folate (p = 0.44, t = 0.39, df = 43) or vitamin B12 (p = 0.21, t = 1.48, df = 43) in the patients in the exacerbation and remission phase of illness. Additionally, there was no significant correlation between serum folate or vitamin B12 levels and scores of positive or negative symptoms in the patient groups. The mean Hcy level was not different between smokers and nonsmokers. In SCH (+) group, smokers (n = 14) had mean Hcy level 15.38 ± 7.10 μmol/L and non-smokers (n = 12) 13.27 ± 5.73 μmol/L (p = 0.703). In the SCH (−) group smokers (n = 11) had a mean Hcy level 12.98 ± 3.8 μmol/L and non-smokers (n = 7) had a mean 11.52 ± 4.73 μmol/L (p = 0.588). There were also no differences in Hcy levels between coffee consumers and non-consumers in either of the SCH patients groups. In the group of coffee consumers (n = 34) the mean Hcy concentration was 13.69 ± 6.59 μmol/L while in the group coffee non-consumers (n = 10) the mean Hcy concentration was 12.30 ± 1.85 μmol/L (p = 0.621). 4. Discussion Elevated plasma Hcy levels have been repeatedly found in young male patients suffering from schizophrenia. Our findings of elevated Hcy levels in young male SCH patients are in agreement with the majority of reported results. Additionally, we found no differences in
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Hcy levels between patients with predominantly negative and predominantly positive schizophrenic symptoms. The origin of elevated plasma Hcy in schizophrenia is unclear, but epidemiological studies suggest that poor nutrition, smoking, coffee consumption and lack of exercise can all contribute to elevated Hcy levels (Schneede et al., 2000). A common genetic polymorphism of methylenetetrahydrofolate reductase (MTHFR) may also predispose individuals to elevated Hcy especially in the presence of the epidemiological risk factors (Cortese and Motti, 2001). Recently, a rigorous meta-analysis was performed by Muntjewerff et al. (2006) which concluded that elevated Hcy levels and the TT genotype of the MTHFR 677C→T polymorphism may contribute to a susceptibility to schizophrenia. While the evidence suggests a possible link between the aberrant MTHFR gene, elevated Hcy, and schizophrenia, this link has not yet been found. We have found a decrease of Hcy levels in the remission phase of schizophrenia compared to the levels in the exacerbation phase. Our data further indicate that the changes are not a consequence of age, duration of illness, folate or vitamin B12 deficiencies, or smoking and coffee consumption. In agreement with our findings, Haidemenos et al. (2007), Regland et al. (1995) and Akanji et al. (2007) found significant elevations of plasma Hcy levels in schizophrenic patients who had folate and vitamin B12 levels within normal limits. Reif et al. (2003) have reported a lack of correlation between Hcy levels and folate and vitamin B12, a finding that is also in agreement with our results. On the other hand, studies have demonstrated reduced folate and vitamin B12 levels in schizophrenic patients (Silver, 2000; Herrán et al., 1999). We have revealed a non-significant decrease in folate concentrations in schizophrenic patients compared to the levels in the control group, although almost all measured values (except one) were in the reference range. It is notable that folate and vitamin B12 concentrations are not changed in our patients after one month of treatment while in this period Hcy levels are normalized. It has been suggested that obesity can also contribute to elevated Hcy levels (Sanchez-Margalet et al., 2002; Konukoğlu et al., 2003). However, several recent studies have failed to detect an association between Hcy levels and BMI (Mora et al., 2006; Papandreou et al., 2007) while Lin et al. (2008) found a significant correlation between Hcy levels and waist-to-hip ratio in male patients with coronary artery disease. Usually elevated plasma Hcy level is considered to be a contributing to the development of pathophysiological changes seen in subjects suffering from schizophrenia. Our findings may indicate that elevated plasma Hcy level may be a consequence of pathological processes involved in schizophrenia. Besides nutrients (mainly folate, vitamins B6 and B12), several factors, including hormones, redox state as well as immune mediators, are involved in the complex regulation of Hcy metabolism (House et al., 1999; Graham and O'Callaghan, 2002; Vitvitsky et al., 2003; Tobena et al., 1996). Stress is one of the factors that could be responsible for changes in Hcy levels in schizophrenic patients. de Oliveira et al. (2004) investigated the effects of stress on total plasma Hcy using four distinct acute stressors in rats: swimming, restrain, novelty and cold exposure. Increased activity of the hypothalamic-pituitary-adrenal (HPA) axis was seen after all stress manipulations except novelty. Plasma Hcy changes occurred in a stressor-specific manner and restrain, which represents a model of psychological stress in rats, significantly increased total plasma Hcy concentration. The influence of stress on the serum Hcy level was also confirmed in a recent investigation by Levine et al. (in press). These authors have found significant elevation of serum Hcy level in male patients with posttraumatic stress disorder (PTSD). The duration of PTSD was found to predict serum Hcy levels. Several studies indicate that a significant number of patients with schizophrenia have elevated baseline plasma cortisol levels that are normalized during the anti-
psychotic treatment (Popovic et al., 2007; Cesková et al., 2006; Cohrs et al., 2006; Meier et al., 2005). Walsh et al. (2005) found higher baseline plasma cortisol and ACTH levels in drug naive patients with schizophrenia. There is mounting evidence for changes in the balance between production and elimination of free radicals in schizophrenic patients (Cadet and Kahler, 1994; Mukerjee et al., 1996; Petronijević et al., 2003). Oxidative stress could represent an endogenous reason for 5,6,7,8-tetrahydrofolate deficiency even when dietary intake of the vitamin is within the recommended range (Widner et al., 2001; Fuchs et al., 2001). Also it has been reported that schizophrenic patients have significantly higher levels of plasma nitrites (Yanik et al., 2003). Nitrous oxide is known to irreversibly inactivate methionine synthase by oxidizing cobalt in methylcobalamin (Danishpajooh et al., 2001). Another enzyme involved in Hcy metabolism, cystathionine βsynthase is also redox-sensitive (Mosharov et al., 2000; Banerjee and Zou, 2005). Growing evidence suggests that neuro-immune-endocrine crosstalk may be impaired in schizophrenia. Cytokine alterations in schizophrenia are currently under intensive investigation and it is suggested that this system may be involved in the neuropathological changes occurring in the central nervous system of schizophrenic patients (Drzyzga et al., 2006). However, there is a lack of data about the influence of cytokines on Hcy metabolism. Only one study has addressed the effects of therapy on Hcy levels in schizophrenic patients. Levine et al. (2006) investigated neuroleptic treated (mean neuroleptic dose in chlorpromazine equivalents was 900 mg; range: 200–1900 mg) patients with plasma Hcy levels above 15 μmol/L in a double-blind crossover trial with the administration of folic acid, pyridoxine, and vitamin B12. The placebo group of patients included in this study received antipsychotic therapy without vitamins and was similar to our groups of patients. However, these authors found a decrease in Hcy levels only in the group of patients with vitamin therapy. They also found an improvement of clinical symptoms of schizophrenia as measured by the Positive and Negative Syndrome Scale with vitamin treatment compared with placebo. These results strongly suggest the benefit of vitamin treatment in SCH patients but do not support our finding of decrease in plasma Hcy concentration in patients subjected to antipsychotic treatment without vitamin therapy. However, there is a significant difference between the placebo group in the aforementioned study and our groups of patients. In the study of Levine et al. (2006) the selected patients had much higher serum Hcy levels at the time of entering the study (mean value in vitamin treated group was around 24 μmol/L and in placebo treated group 28 μmol/L) than those included in our groups (that are usually observed in schizophrenic patients). Since only a subgroup of schizophrenic patients with extremely high Hcy levels was included, it is possible that this selection recruits the subjects with some inherited defect in enzymes involved in Hcy metabolism. The finding of a positive correlation between the score of negative symptoms and Hcy concentration in our patients is interesting, especially since it is found in both SCH (+) and SCH (−) groups of patients. This could indicate the influence of a pathological process responsible for development of negative symptoms on Hcy levels. There are some studies that put the light on the relationship between score of negative symptoms and Hcy metabolism in schizophrenia. Goff et al. (2004) have found that folate concentrations inversely correlate with scores on the Scale for Assessment of Negative Symptoms in nonsmokers; however, Hcy concentration remained unchanged in comparison to the control group. Roffman et al. (2008) have found that methylenetetrahydrofolate reductase polymorphism contributes to variation in negative symptom severity in schizophrenia although serum folate did not differ significantly among the genotype groups studied. Finally, several limitations should be mentioned for the interpretation of our results. Only male patients were included and all
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patients were medicated. We did not assess body mass index, lipid status or methylenetetrahydrofolate reductase polymorphism. 5. Conclusion We can conclude that young male schizophrenic patients have increased plasma Hcy levels during the exacerbation phase of the illness, regardless of age, duration of illness, folate and vitamin B12 deficiencies, or smoking and coffee consumption. This elevation was seen both in patients with a predominantly positive and predominantly negative syndrome in schizophrenia. One month later, in the remission phase of the illness, Hcy concentrations were significantly decreased while serum folate and vitamin B12 concentrations remained unchanged in comparison to the values in the exacerbation phase. Further experiments are needed to clarify the interrelationship between Hcy and schizophrenia. Acknowledgements We thank Urosh Vilimanovich for the help in editing this manuscript. This study was supported by grant # 145058 from the Ministry for Science and Environmental Protection of the Republic of Serbia. References Adler Nevo G, Meged S, Sela BA, Hanoch-Levi A, Hershko R, Weizman A. Homocysteine levels in adolescent schizophrenia patients. Eur Neuropsychopharmacol 2006;16: 588–91. Akanji AO, Ohaeri JU, Al-Shammri SA, Fatania HR. Associations of blood homocysteine concentrations in Arab schizophrenic patients. Clin Biochem 2007;40:1026–31. American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders (DSM-IV). 4th ed. Washington, DC: American Psychiatric Association; 1994. Amin F, Silverman JM, Siever LJ, Smith CJ, Knott PJ, Davis KL. Genetic antecedents of dopamine dysfunction in schizophrenia. Biol Psychiatry 1999;45:1143–50. Applebaum J, Shimon H, Sela BA, Belmaker RH, Levine J. Homocysteine levels in newly admitted schizophrenic patients. J Psychiatr Res 2004;38:413–6. Banerjee R, Zou CG. Redox regulation and reaction mechanism of human cystathioninebeta-synthase: a PLP-dependent hemesensor protein. Arch Biochem Biophys 2005;433: 144–56. Bottiglieri T, Laundy M, Crellin R, Toone BK, Carney MW, Reynolds EH. Homocysteine, folate, methylation, and monoamine metabolism in depression. J Neurol Neurosurg Psychiatry 2000;69:228–32. Brown AS, Bottiglieri T, Schaefer CA, Quesenberry Jr CP, Liu L, Bresnahan M, et al. Elevated prenatal homocysteine levels as a risk factor for schizophrenia. Arch Gen Psychiatry 2007;64:31–9. Cadet JL, Kahler LA. Free radical mechanisms in schizophrenia and tardive dyskinesia. Neurosci Biobehav Rev 1994;18:457–67. Cesková E, Kaspárek T, Zourková A, Prikryl R. Dexamethasone suppression test in firstepisode schizophrenia. Neuro Endocrinol Lett 2006;27:433–7. Cohrs S, Röher C, Jordan W, Meier A, Huether G, Wuttke W, et al. The atypical antipsychotics olanzapine and quetiapine, but not haloperidol, reduce ACTH and cortisol secretion in healthy subjects. Psychopharmacology (Berl) 2006;185:11–8. Cortese C, Motti C. MTHFR gene polymorphism, homocysteine and cardiovascular disease. Public Health Nutr 2001;4:493–7. Danishpajooh IO, Gudi T, Chen Y, Kharitonov VG, Sharma VS, Boss GR. Nitric oxide inhibits methionine synthase activity in vivo and disrupts carbon flow through the folate pathway. J Biol Chem 2001;276:27296–303. de Oliveira AC, Suchecki D, Cohen S, D'Almeida V. Acute stressor-selective effect on total plasma homocysteine concentration in rats. Pharmacol Biochem Behav 2004;77: 269–73. Drzyzga L, Obuchowicz E, Marcinowska A, Herman ZS. Cytokines in schizophrenia and the effects of antipsychotic drugs. Brain Behav Immun 2006;20:532–45. Folstein M, Liu T, Peter I, Buell J, Arsenault L, Scott T, et al. The homocysteine hypothesis of depression. Am J Psychiatry 2007;164:861–7 Review. Fuchs D, Jaeger M, Widner B, Wirleitner B, Artner-Dworzak E, Leblhuber F. Is hyperhomocysteinemia due to the oxidative depletion of folate rather than to insufficient dietary intake? Clin Chem Lab Med 2001;39:691–4. Furie KL, Kelly PJ. Homocyst(e)ine and stroke. Semin Neurol 2006;26:24–32 Review. Graham IM, O'Callaghan P. Vitamins, homocysteine and cardiovascular risk. Cardiovasc Drugs Ther 2002;16:383–9. Goff DC, Bottiglieri T, Arning E, Shih V, Freudenreich O, Evins AE, et al. Folate, homocysteine, and negative symptoms in schizophrenia. Am J Psychiatry 2004;161:1705–8. Haidemenos A, Kontis D, Gazi A, Kallai E, Allin M, Lucia B. Plasma homocysteine, folate and B12 in chronic schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry 2007;31:1289–96. Hassin-Baer S, Cohen O, Vakil E, Sela BA, Nitsan Z, Schwartz R, et al. Plasma homocysteine levels and Parkinson disease: disease progression, carotid intima-media thickness and neuropsychiatric complications. Clin Neuropharmacol 2006;29:305–11.
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Progress in Neuro-Psychopharmacology & Biological Psychiatry j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / p n p b p
Plasma homocysteine levels in young male patients in the exacerbation and remission phase of schizophrenia Nataša D. Petronijević a,⁎, Nevena V. Radonjić a, Maja D. Ivković b, Dragan Marinković b, Vesna D. Piperski c, Bogdan M. Đuričić a, Vladimir R. Paunović b a b c
Institute of Medical and Clinical Biochemistry, School of Medicine, Pasterova 2, 11000 Belgrade, Serbia Institute of Psychiatry, School of Medicine, Pasterova 2, 11000 Belgrade, Serbia Present address: Medical Academy US Medical School, Kumodraška 261, 11000 Beograd, Serbia
a r t i c l e
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Article history: Received 17 March 2008 Received in revised form 10 September 2008 Accepted 11 September 2008 Available online 15 September 2008 Keywords: B12 Folate Homocysteine Negative symptoms Positive symptoms Schizophrenia
a b s t r a c t High levels of homocysteine (Hcy) were suggested to contribute to the pathogenesis of schizophrenia. Recent investigations have shown that treatment with folic acid, vitamin B-12 and pyridoxine are effective in reducing Hcy levels while concomitantly reducing the score of positive and negative symptoms in schizophrenic patients. In addition to the availability of nutrients (mainly folate, vitamins B6 and B12), plasma Hcy concentrations are dependent on complex metabolic regulation that could be disrupted in schizophrenia. This study was designed to test the influence of disease activity on plasma Hcy levels. Plasma Hcy concentrations were measured in male chronic schizophrenic patients with a predominantly positive (SCH (+)) or predominantly negative (SCH (−)) syndrome in schizophrenia immediately upon admission to the hospital (exacerbation phase) and one month later (remission phase). During this period patients received antipsychotic medications without vitamin therapy. The effects of age, duration of illness, folate and B12 concentrations, as well as smoking and coffee consumption habits on the observed changes were evaluated. Age- and sex-matched subjects were included in the control group. In the control group plasma Hcy concentration was 8.75 ± 1.84 μmol/L. In the exacerbation phase plasma Hcy concentrations were significantly increased both in SCH (+) (14.91 ± 6.19 μmol/L) and SCH (−) groups (12.8 ± 3.27 μmol/L). There was no difference in plasma Hcy concentrations between SCH (+) and SCH (−) patients. Serum folate and B12 concentrations were not significantly different in any of the investigated groups of subjects. The plasma Hcy concentrations could not be correlated with age, duration of illness, the score of positive symptoms or the concentration of folate and vitamin B12. A positive correlation was found between plasma Hcy level and score of negative symptoms in both groups of patients. No correlation was found between smoking or coffee consumption habits and plasma Hcy concentrations. All patients exhibited decreased plasma Hcy levels in the remission phase of the illness, with a mean decrease of 2.68 ± 1.57 μmol/L. Folate and B12 levels did not differ in the exacerbation and remission phases of the illness. The significant decrease of plasma Hcy levels, without changes in folate and vitamin B12 concentrations in the remission phase of schizophrenia, could indicate an influence of a pathogenetic process involved in schizophrenia on Hcy metabolism. © 2008 Elsevier Inc. All rights reserved.
1. Introduction Homocysteine (Hcy) is a sulfur containing amino acid formed during the metabolism of the essential amino acid methionine. In most tissues, Hcy may be remethylated to methionine by the enzyme
Abbreviations: BMI, body mass index; EDTA, ethylenediaminetetraacetic acid; EIA, enzyme immunoassay; Hcy, homocysteine; HPA, hypothalamic-pituitary-adrenal; MTHFR, methylenetetrahydrofolate reductase; PANSS, positive and negative syndrome scale; PTSD, posttraumatic stress disorder; SCH, schizophrenia. ⁎ Corresponding author. Tel./fax: +381 11 2682 953. E-mail address: [email protected] (N.D. Petronijević). 0278-5846/$ – see front matter © 2008 Elsevier Inc. All rights reserved. doi:10.1016/j.pnpbp.2008.09.009
methionine synthase by a reaction where vitamin B12 is a cofactor and methyltetrahydrofolate is a substrate. Elevated level of plasma total Hcy was found to be a risk factor for cardiovascular (Nygård et al., 1995) and cerebrovascular disease (Furie and Kelly, 2006). Increased total plasma Hcy levels have also been reported in several neuropsychiatric disorders including Alzheimer's disease (Seshadri et al., 2002), Parkinson's disease (O'Suilleabhain et al., 2006; Hassin-Baer et al., 2006; Todorović et al., 2006), depression (Bottiglieri et al., 2000; Folstein et al., 2007), bipolar disorder (Osher et al., 2004), schizophrenia and schizophrenia-like psychosis (Regland et al., 1995; Susser et al., 1998; Levine et al., 2002). Markedly elevated plasma Hcy levels have been found in young male schizophrenic patients (Levine et al., 2002), a finding that has
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been replicated in a group of newly admitted schizophrenic patients (Applebaum et al., 2004), as well as in adolescents (age 14–21 years) suffering from schizophrenia (Adler Nevo et al., 2006). On the other hand, reports regarding hyperhomocysteinemia in female schizophrenic patients have not been consistent. Haidemenos et al. (2007) did not find gender and age differences of plasma Hcy levels in schizophrenic patients, suggesting that hyperhomocystinemia may be a risk factor for schizophrenia regardless of gender. However, another study failed to detect elevated Hcy levels in young schizophrenic women (Reif et al., 2003). Notably, elevated Hcy levels among pregnant women have been found to increase the risk of their children developing schizophrenia (Brown et al., 2007). High levels of Hcy have been suggested to contribute to the pathogenesis of schizophrenia (Levine et al., 2005). Recently, studies have suggested that reducing Hcy levels is an effective strategy to ameliorate clinical symptoms in schizophrenic patients. Levine et al. (2006) has shown that treatment with folic acid, vitamin B-12 and pyridoxine is effective in reducing Hcy levels as well as the scores of positive and negative symptoms in schizophrenic patients with starting plasma Hcy concentrations above 15 μmol/L. The results of a preliminary study by Miodownik et al. (2007) indicated the effectiveness of high dose vitamin B6 in decreasing serum Hcy levels in male schizophrenic patients. The rationale for the current study was to determine whether the elevated homocysteine level reported in this disorder is a causative factor or is a result of a pathophysiologic process of schizophrenia and if such an elevated level is a state or a trait marker for this disease. Plasma Hcy levels were measured in young male chronic schizophrenic patients with predominantly positive or predominantly negative symptoms, during the exacerbation phase of the illness and one month later, in the remission phase. During this period patients received neuroleptic medication without vitamin supplements. The use of different antipsychotics showed no trend for association with increased Hcy (Levine et al., 2005). The effects of age, duration of illness, folate and vitamin B12 concentrations, as well as smoking and coffee consumption habits, on observed changes were evaluated. 2. Patients and methods Fifty-two male patients from the Institute of Psychiatry, School of Medicine, University of Belgrade, which met DSM-IV criteria (American Psychiatric Association, 1994) for schizophrenia and 20 healthy control male healthy volunteers recruited from the staff of the Institute of Psychiatry and Institute of Medical and Clinical Biochemistry were included in the study. Three of the recruited patients did not agree to participate in the study and five others were not in the remission phase after one month. The final patient group consisted of 44 subjects. In the SCH group, psychopathology was assessed by Positive and Negative Syndrome Scale (PANSS). All patients were divided into two groups: with either a predominantly positive [SCH (+)] or a predominantly negative [SCH (−)] syndrome in schizophrenia (SCH) (Kay et al., 1987). Empirically established percentile ranks derived from raw and standardized T-scores on the PANSS profile form were used to suit research objectives. This procedure declares that a person with decidedly high positive or negative profile scores is at the extremes of the schizophrenic sample-distribution curve for the PANSS composite scale. The authors used the most rigorous criterion. Consequently, only those patients ranked above the 95th percentile (composite score of 11 and higher) were considered SCH (+), whereas those ranking below the 25th percentile (composite scores of −15 and lower) were considered SCH (−). All subjects were otherwise medically healthy and no subjects had histories of cardiovascular or neurological disease, diabetes, or a history of substance abuse or dependence. Written informed consent was obtained from each subject after a complete description of the study was provided. Patients were introduced into the study in the exacerbation phase. Twenty-six SCH patients had a predominantly positive (21–
50 years old; mean 35.28 ± 8.28) and eighteen a predominantly negative (21–50 years old; mean 33.78 ± 9.37) syndrome. All patients included in the study were treated with antipsychotic drugs (mean neuroleptic dose in chlorpromazine equivalents was 276 ± 161 mg per day; range: 100–630 mg per day). Patients were also grouped as smokers or non-smokers (smokers consumed 16.2 ± 3.5 cigarettes per day) and according to their coffee consumption habits (coffee consumers drink at least one cup of coffee per day). The blood samples were collected immediately upon admission to the hospital in the exacerbation phase and again one month later in the remission phase of illness. All biochemical measurements were conducted blind to diagnostic status. During the study all patients were on regular hospital diet, and there were no clinically relevant symptoms of malnutrition. Control subjects (n = 20) were male healthy volunteers (25– 45 years old; mean 33 ± 8.1). Because non-psychotic, first-degree relatives of SCH probands may carry a genetic diathesis to SCH (Amin et al., 1999), the subjects included in the control group were required to be free of any major psychiatric disorder in their first-degree relatives. Additionally, they were required to have a negative family history of dementia, mental retardation, and Huntington's disease in first-degree relatives. Smoking was prohibited after 11 p.m. before the next day's blood draw. None of the subjects had taken any vitamin supplements three months prior to being included in the study. Subjects also did not take any medications (carbamazepine, phenytoin, anticonvulsants or 6-azauridine triacetate) that are known to cause an elevation of plasma Hcy levels. Food consumption can affect circulating Hcy levels and protein rich meals were avoided late in the day before blood sampling. 2.1. Collection of blood samples and biochemical analyses Blood samples were taken from antecubital vein between 7 and 8 a.m. into a vacutainer containing 1 mL 1% EDTA as an anticoagulant. The samples were kept on ice (maximum of 1 h) and plasma and blood cells were separated by centrifugation (15 min, 3000 rpm). Plasma samples were stored at −80 °C until Hcy measurements. Hcy levels were measured by Bio-Rad Microplate Enzyme Immunoassay Homocysteine Test (EIA) (Bio-Rad Laboratories, Inc., Hercules, CA, USA). Serum levels of folate and vitamin B12 were determined by competitive chemiluminescent assays with an IMMULITE 1000 analyzer (Siemens Healthcare Diagnostics, NY, USA). The data sets were tested for normal distribution by the Kolmogorov– Smirnov test. All data sets were within normal distribution and analyzed using the analysis of variance (ANOVA) followed by Fisher's test for multiple comparison. Associations between age, duration of illness, clinical rating scale scores, serum folate and B12 and levels of plasma Hcy were tested by Pearson's correlation coefficient. For assessment of
Fig. 1. Plasma Hcy levels in control subjects (n = 20) and patients suffering from schizophrenia included in SCH (+) (n = 26) and SCH (−) (n = 18) group. ⁎p b 0.01 in SCH (+) group vs. controls and p b 0.05 in SCH (−) group vs. controls.
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Fig. 3. Plasma Hcy concentration in all SCH patients in the exacerbation phase upon admission and in the remission phase of illness one month later.
Fig. 2. Correlation of plasma Hcy levels with the score of negative (A and B) symptoms in SCH (+) (n = 26) and SCH (−) (n = 18) patients.
smoking or coffee consumption influence on the plasma Hcy levels in patients with predominantly positive or negative syndrome in SCH, the Student's t-test was used. Significance of plasma Hcy and serum folate and vitamin B12 concentration changes in exacerbation and remission phase of illness was tested with paired t-test. Results are presented as mean ±SD and pb 0.05 was regarded as significant for all comparisons. 3. Results When compared to healthy controls, plasma Hcy concentrations (Fig. 1) were significantly elevated in both SCH (+) and SCH (−) patients immediately upon admission to the hospital (df = 2; p = 0.0001). Although SCH (+) patients had higher plasma Hcy concentrations than SCH (−) patients, the difference was not statistically significant. Statistically significant correlations between plasma Hcy concentrations and age (R = 0.17, p = 0.65 for SCH (+); R = 0.48, p = 0.19 for SCH (−)) or duration of illness (R = 0.37, p = 0.24 for SCH (+); R = 0.51, p = 0.25 for SCH (−)) were not found in either SCH (+) or SCH (−) patients. Correlations of plasma Hcy levels and the scores of negative symptoms are presented on Fig. 2. A significant correlation was found
Table 1 Serum folate and vitamin B12 levels in control group and SCH (+) and SCH (−) groups of patients immediately upon admission to the hospital Controls
SCH (+)
SCH (−)
n = 20
n = 26
n = 18
SCH (+) vs. SCH (−) vs. SCH (+) vs. control control SCH (−)
Folate (ng/mL) 8.19 ± 1.69 6.35 ± 1.69 7.67 ± 1.89 p = 0.08 B12 (pg/mL) 397 ± 241 516 ± 296 363 ± 163 p = 0.45
p = 0.72 p = 0.61
Values are represented as mean ± SD for specified number of subjects.
p = 0.21 p = 0.32
between plasma Hcy levels and scores of negative symptoms in SCH (+) as well as in SCH (−) group (Fig. 2). At the same time there was no significant correlation between plasma Hcy levels and scores of positive symptoms in both SCH (+) (R = 0.28, p = 0.166) and SCH (−) (R = −0.195, p = 0.43) groups. The concentrations of serum folate and vitamin B12 did not differ between investigated groups of subjects (Table 1). Although the concentration of folate was lower in the SCH (+) group compared to the control group, the difference did not reach statistical significance. There was no significant correlation between plasma Hcy and folate (R = − 0.317, df = 43, p = 0.14) or vitamin B12 (R = − 0.314, df = 43, p = 0.154) levels in patients upon admission to the hospital. Comparison of plasma Hcy levels measured immediately upon admission to the hospital and one month later is presented at Fig. 3. The mean change of plasma Hcy level between those two measurements was 2.68 ± 1.57 μmol/L, and the difference was statistically significant (p = 0.00478: t = 10.42; df = 43). Regardless of the starting Hcy level, decreased plasma Hcy levels were observed in every patient tested. In the remission phase mean plasma Hcy level in SCH (+) group was 11.18 ± 4.7 μmol/L (p = 0.14 comparing to control group) and in SCH (−) group 10.07 ± 2.51 μmol/L (p = 0.21 comparing to control group), suggesting that after one month of treatment, regardless of positive or negative symptomatology upon admission, patient plasma Hcy levels were indistinguishable from those of the control group. The levels of folate and vitamin B12 did not differ between patients immediately after admission to the hospital. As investigated by paired t-test, there were no significant differences in the between levels of folate (p = 0.44, t = 0.39, df = 43) or vitamin B12 (p = 0.21, t = 1.48, df = 43) in the patients in the exacerbation and remission phase of illness. Additionally, there was no significant correlation between serum folate or vitamin B12 levels and scores of positive or negative symptoms in the patient groups. The mean Hcy level was not different between smokers and nonsmokers. In SCH (+) group, smokers (n = 14) had mean Hcy level 15.38 ± 7.10 μmol/L and non-smokers (n = 12) 13.27 ± 5.73 μmol/L (p = 0.703). In the SCH (−) group smokers (n = 11) had a mean Hcy level 12.98 ± 3.8 μmol/L and non-smokers (n = 7) had a mean 11.52 ± 4.73 μmol/L (p = 0.588). There were also no differences in Hcy levels between coffee consumers and non-consumers in either of the SCH patients groups. In the group of coffee consumers (n = 34) the mean Hcy concentration was 13.69 ± 6.59 μmol/L while in the group coffee non-consumers (n = 10) the mean Hcy concentration was 12.30 ± 1.85 μmol/L (p = 0.621). 4. Discussion Elevated plasma Hcy levels have been repeatedly found in young male patients suffering from schizophrenia. Our findings of elevated Hcy levels in young male SCH patients are in agreement with the majority of reported results. Additionally, we found no differences in
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Hcy levels between patients with predominantly negative and predominantly positive schizophrenic symptoms. The origin of elevated plasma Hcy in schizophrenia is unclear, but epidemiological studies suggest that poor nutrition, smoking, coffee consumption and lack of exercise can all contribute to elevated Hcy levels (Schneede et al., 2000). A common genetic polymorphism of methylenetetrahydrofolate reductase (MTHFR) may also predispose individuals to elevated Hcy especially in the presence of the epidemiological risk factors (Cortese and Motti, 2001). Recently, a rigorous meta-analysis was performed by Muntjewerff et al. (2006) which concluded that elevated Hcy levels and the TT genotype of the MTHFR 677C→T polymorphism may contribute to a susceptibility to schizophrenia. While the evidence suggests a possible link between the aberrant MTHFR gene, elevated Hcy, and schizophrenia, this link has not yet been found. We have found a decrease of Hcy levels in the remission phase of schizophrenia compared to the levels in the exacerbation phase. Our data further indicate that the changes are not a consequence of age, duration of illness, folate or vitamin B12 deficiencies, or smoking and coffee consumption. In agreement with our findings, Haidemenos et al. (2007), Regland et al. (1995) and Akanji et al. (2007) found significant elevations of plasma Hcy levels in schizophrenic patients who had folate and vitamin B12 levels within normal limits. Reif et al. (2003) have reported a lack of correlation between Hcy levels and folate and vitamin B12, a finding that is also in agreement with our results. On the other hand, studies have demonstrated reduced folate and vitamin B12 levels in schizophrenic patients (Silver, 2000; Herrán et al., 1999). We have revealed a non-significant decrease in folate concentrations in schizophrenic patients compared to the levels in the control group, although almost all measured values (except one) were in the reference range. It is notable that folate and vitamin B12 concentrations are not changed in our patients after one month of treatment while in this period Hcy levels are normalized. It has been suggested that obesity can also contribute to elevated Hcy levels (Sanchez-Margalet et al., 2002; Konukoğlu et al., 2003). However, several recent studies have failed to detect an association between Hcy levels and BMI (Mora et al., 2006; Papandreou et al., 2007) while Lin et al. (2008) found a significant correlation between Hcy levels and waist-to-hip ratio in male patients with coronary artery disease. Usually elevated plasma Hcy level is considered to be a contributing to the development of pathophysiological changes seen in subjects suffering from schizophrenia. Our findings may indicate that elevated plasma Hcy level may be a consequence of pathological processes involved in schizophrenia. Besides nutrients (mainly folate, vitamins B6 and B12), several factors, including hormones, redox state as well as immune mediators, are involved in the complex regulation of Hcy metabolism (House et al., 1999; Graham and O'Callaghan, 2002; Vitvitsky et al., 2003; Tobena et al., 1996). Stress is one of the factors that could be responsible for changes in Hcy levels in schizophrenic patients. de Oliveira et al. (2004) investigated the effects of stress on total plasma Hcy using four distinct acute stressors in rats: swimming, restrain, novelty and cold exposure. Increased activity of the hypothalamic-pituitary-adrenal (HPA) axis was seen after all stress manipulations except novelty. Plasma Hcy changes occurred in a stressor-specific manner and restrain, which represents a model of psychological stress in rats, significantly increased total plasma Hcy concentration. The influence of stress on the serum Hcy level was also confirmed in a recent investigation by Levine et al. (in press). These authors have found significant elevation of serum Hcy level in male patients with posttraumatic stress disorder (PTSD). The duration of PTSD was found to predict serum Hcy levels. Several studies indicate that a significant number of patients with schizophrenia have elevated baseline plasma cortisol levels that are normalized during the anti-
psychotic treatment (Popovic et al., 2007; Cesková et al., 2006; Cohrs et al., 2006; Meier et al., 2005). Walsh et al. (2005) found higher baseline plasma cortisol and ACTH levels in drug naive patients with schizophrenia. There is mounting evidence for changes in the balance between production and elimination of free radicals in schizophrenic patients (Cadet and Kahler, 1994; Mukerjee et al., 1996; Petronijević et al., 2003). Oxidative stress could represent an endogenous reason for 5,6,7,8-tetrahydrofolate deficiency even when dietary intake of the vitamin is within the recommended range (Widner et al., 2001; Fuchs et al., 2001). Also it has been reported that schizophrenic patients have significantly higher levels of plasma nitrites (Yanik et al., 2003). Nitrous oxide is known to irreversibly inactivate methionine synthase by oxidizing cobalt in methylcobalamin (Danishpajooh et al., 2001). Another enzyme involved in Hcy metabolism, cystathionine βsynthase is also redox-sensitive (Mosharov et al., 2000; Banerjee and Zou, 2005). Growing evidence suggests that neuro-immune-endocrine crosstalk may be impaired in schizophrenia. Cytokine alterations in schizophrenia are currently under intensive investigation and it is suggested that this system may be involved in the neuropathological changes occurring in the central nervous system of schizophrenic patients (Drzyzga et al., 2006). However, there is a lack of data about the influence of cytokines on Hcy metabolism. Only one study has addressed the effects of therapy on Hcy levels in schizophrenic patients. Levine et al. (2006) investigated neuroleptic treated (mean neuroleptic dose in chlorpromazine equivalents was 900 mg; range: 200–1900 mg) patients with plasma Hcy levels above 15 μmol/L in a double-blind crossover trial with the administration of folic acid, pyridoxine, and vitamin B12. The placebo group of patients included in this study received antipsychotic therapy without vitamins and was similar to our groups of patients. However, these authors found a decrease in Hcy levels only in the group of patients with vitamin therapy. They also found an improvement of clinical symptoms of schizophrenia as measured by the Positive and Negative Syndrome Scale with vitamin treatment compared with placebo. These results strongly suggest the benefit of vitamin treatment in SCH patients but do not support our finding of decrease in plasma Hcy concentration in patients subjected to antipsychotic treatment without vitamin therapy. However, there is a significant difference between the placebo group in the aforementioned study and our groups of patients. In the study of Levine et al. (2006) the selected patients had much higher serum Hcy levels at the time of entering the study (mean value in vitamin treated group was around 24 μmol/L and in placebo treated group 28 μmol/L) than those included in our groups (that are usually observed in schizophrenic patients). Since only a subgroup of schizophrenic patients with extremely high Hcy levels was included, it is possible that this selection recruits the subjects with some inherited defect in enzymes involved in Hcy metabolism. The finding of a positive correlation between the score of negative symptoms and Hcy concentration in our patients is interesting, especially since it is found in both SCH (+) and SCH (−) groups of patients. This could indicate the influence of a pathological process responsible for development of negative symptoms on Hcy levels. There are some studies that put the light on the relationship between score of negative symptoms and Hcy metabolism in schizophrenia. Goff et al. (2004) have found that folate concentrations inversely correlate with scores on the Scale for Assessment of Negative Symptoms in nonsmokers; however, Hcy concentration remained unchanged in comparison to the control group. Roffman et al. (2008) have found that methylenetetrahydrofolate reductase polymorphism contributes to variation in negative symptom severity in schizophrenia although serum folate did not differ significantly among the genotype groups studied. Finally, several limitations should be mentioned for the interpretation of our results. Only male patients were included and all
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patients were medicated. We did not assess body mass index, lipid status or methylenetetrahydrofolate reductase polymorphism. 5. Conclusion We can conclude that young male schizophrenic patients have increased plasma Hcy levels during the exacerbation phase of the illness, regardless of age, duration of illness, folate and vitamin B12 deficiencies, or smoking and coffee consumption. This elevation was seen both in patients with a predominantly positive and predominantly negative syndrome in schizophrenia. One month later, in the remission phase of the illness, Hcy concentrations were significantly decreased while serum folate and vitamin B12 concentrations remained unchanged in comparison to the values in the exacerbation phase. Further experiments are needed to clarify the interrelationship between Hcy and schizophrenia. Acknowledgements We thank Urosh Vilimanovich for the help in editing this manuscript. This study was supported by grant # 145058 from the Ministry for Science and Environmental Protection of the Republic of Serbia. References Adler Nevo G, Meged S, Sela BA, Hanoch-Levi A, Hershko R, Weizman A. Homocysteine levels in adolescent schizophrenia patients. Eur Neuropsychopharmacol 2006;16: 588–91. Akanji AO, Ohaeri JU, Al-Shammri SA, Fatania HR. Associations of blood homocysteine concentrations in Arab schizophrenic patients. Clin Biochem 2007;40:1026–31. American Psychiatric Association. Diagnostic and Statistical Manual of Mental Disorders (DSM-IV). 4th ed. Washington, DC: American Psychiatric Association; 1994. Amin F, Silverman JM, Siever LJ, Smith CJ, Knott PJ, Davis KL. Genetic antecedents of dopamine dysfunction in schizophrenia. Biol Psychiatry 1999;45:1143–50. Applebaum J, Shimon H, Sela BA, Belmaker RH, Levine J. Homocysteine levels in newly admitted schizophrenic patients. J Psychiatr Res 2004;38:413–6. Banerjee R, Zou CG. Redox regulation and reaction mechanism of human cystathioninebeta-synthase: a PLP-dependent hemesensor protein. Arch Biochem Biophys 2005;433: 144–56. Bottiglieri T, Laundy M, Crellin R, Toone BK, Carney MW, Reynolds EH. Homocysteine, folate, methylation, and monoamine metabolism in depression. J Neurol Neurosurg Psychiatry 2000;69:228–32. Brown AS, Bottiglieri T, Schaefer CA, Quesenberry Jr CP, Liu L, Bresnahan M, et al. Elevated prenatal homocysteine levels as a risk factor for schizophrenia. Arch Gen Psychiatry 2007;64:31–9. Cadet JL, Kahler LA. Free radical mechanisms in schizophrenia and tardive dyskinesia. Neurosci Biobehav Rev 1994;18:457–67. Cesková E, Kaspárek T, Zourková A, Prikryl R. Dexamethasone suppression test in firstepisode schizophrenia. Neuro Endocrinol Lett 2006;27:433–7. Cohrs S, Röher C, Jordan W, Meier A, Huether G, Wuttke W, et al. The atypical antipsychotics olanzapine and quetiapine, but not haloperidol, reduce ACTH and cortisol secretion in healthy subjects. Psychopharmacology (Berl) 2006;185:11–8. Cortese C, Motti C. MTHFR gene polymorphism, homocysteine and cardiovascular disease. Public Health Nutr 2001;4:493–7. Danishpajooh IO, Gudi T, Chen Y, Kharitonov VG, Sharma VS, Boss GR. Nitric oxide inhibits methionine synthase activity in vivo and disrupts carbon flow through the folate pathway. J Biol Chem 2001;276:27296–303. de Oliveira AC, Suchecki D, Cohen S, D'Almeida V. Acute stressor-selective effect on total plasma homocysteine concentration in rats. Pharmacol Biochem Behav 2004;77: 269–73. Drzyzga L, Obuchowicz E, Marcinowska A, Herman ZS. Cytokines in schizophrenia and the effects of antipsychotic drugs. Brain Behav Immun 2006;20:532–45. Folstein M, Liu T, Peter I, Buell J, Arsenault L, Scott T, et al. The homocysteine hypothesis of depression. Am J Psychiatry 2007;164:861–7 Review. Fuchs D, Jaeger M, Widner B, Wirleitner B, Artner-Dworzak E, Leblhuber F. Is hyperhomocysteinemia due to the oxidative depletion of folate rather than to insufficient dietary intake? Clin Chem Lab Med 2001;39:691–4. Furie KL, Kelly PJ. Homocyst(e)ine and stroke. Semin Neurol 2006;26:24–32 Review. Graham IM, O'Callaghan P. Vitamins, homocysteine and cardiovascular risk. Cardiovasc Drugs Ther 2002;16:383–9. Goff DC, Bottiglieri T, Arning E, Shih V, Freudenreich O, Evins AE, et al. Folate, homocysteine, and negative symptoms in schizophrenia. Am J Psychiatry 2004;161:1705–8. Haidemenos A, Kontis D, Gazi A, Kallai E, Allin M, Lucia B. Plasma homocysteine, folate and B12 in chronic schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry 2007;31:1289–96. Hassin-Baer S, Cohen O, Vakil E, Sela BA, Nitsan Z, Schwartz R, et al. Plasma homocysteine levels and Parkinson disease: disease progression, carotid intima-media thickness and neuropsychiatric complications. Clin Neuropharmacol 2006;29:305–11.
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