Plasma antioxidant capacity is reduced in Asperger syndrome

Journal of Psychiatric Research 46 (2012) 394e401

Contents lists available at SciVerse ScienceDirect

Journal of Psychiatric Research journal homepage: www.elsevier.com/locate/psychires

Plasma antioxidant capacity is reduced in Asperger syndrome Mara Parellada a, *, Carmen Moreno a, Karina Mac-Dowell b, c, Juan Carlos Leza b, c, Marisa Giraldez b, Concepción Bailón d, Carmen Castro e, Patricia Miranda-Azpiazu f, David Fraguas g, Celso Arango a a

Child and Adolescent Psychiatry, Department of Psychiatry, Hospital General Universitario Gregorio Marañón, Centro de Investigación en Red de Salud Mental, CIBERSAM, Dr Esquerdo 46, Madrid, Spain b Centro de Investigación Biomédica en Red de Salud Mental, CIBERSAM, Dr Esquerdo 46, 28007, Madrid, Spain c Dept. of Pharmacology, School of Medicine, Univ. Complutense, and Instituto de Investigación Sanitaria iþ12, Hospital Universitario 12 de Octubre, Ciudad Universitaria 28040 Madrid, Spain d Child and Adolescent Psychiatry, Department of Psychiatry, Hospital General Universitario Gregorio Marañón, Ibiza 43, Madrid, 28009, Spain e Division of Physiology, University of Cádiz; Plaza de Falla 9, 11003 Cádiz, Spain f Department of Neurosciences, Pharmacology & Psychiatry. Centro de Investigación Biomédica en Red de Salud Mental, CIBERSAM, Plaza de Falla 9, 11003 Cádiz, Spain g Department of Mental Health, Complejo Hospitalario Universitario de Albacete, Hermanos Falcó 37, 02006 Albacete, Spain

a r t i c l e i n f o

a b s t r a c t

Article history: Received 5 July 2011 Received in revised form 7 October 2011 Accepted 13 October 2011

Recent evidence suggests that children with autism have impaired detoxification capacity and may suffer from chronic oxidative stress. To our knowledge, there has been no study focusing on oxidative metabolism specifically in Asperger syndrome (a milder form of autism) or comparing this metabolism with other psychiatric disorders. In this study, total antioxidant status (TAOS), non-enzymatic (glutathione and homocysteine) and enzymatic (catalase, superoxide dismutase, and glutathione peroxidase) antioxidants, and lipid peroxidation were measured in plasma or erythrocyte lysates in a group of adolescent patients with Asperger syndrome, a group of adolescents with a first episode of psychosis, and a group of healthy controls at baseline and at 8e12 weeks. TAOS was also analyzed at 1 year. TAOS was reduced in Asperger individuals compared with healthy controls and psychosis patients, after covarying by age and antipsychotic treatment. This reduced antioxidant capacity did not depend on any of the individual antioxidant variables measured. Psychosis patients had increased homocysteine levels in plasma and decreased copper and ceruloplasmin at baseline. In conclusion, Asperger patients seem to have chronic low detoxifying capacity. No impaired detoxifying capacity was found in the first-episode psychosis group in the first year of illness. Ó 2011 Elsevier Ltd. All rights reserved.

Keywords: Psychosis Autism spectrum disorders Oxidative metabolism Hyperhomocysteine Redox system

1. Introduction An adequate equilibrium between the production of reactive oxygen species (ROS) and antioxidant capacity plays an important role in cell physiology and brain development. An excessive prooxidant cell environment produces a series of chain reactions that may damage proteins, lipids, and nucleic acids (McCord, 2000; Adibhatla and Hatcher, 2010). Particularly in the brain, an

* Corresponding author. Department of Psychiatry, Hospital General Universitario Gregorio Marañón, Ibiza 43, 28009 Madrid, Spain. Tel.: þ34 91 5868133; fax: þ34 91 4265004. E-mail addresses: [email protected] (M. Parellada), [email protected] (C. Moreno), [email protected] (K. Mac-Dowell), [email protected] (J.C. Leza), [email protected] (M. Giraldez), [email protected] (C. Bailón), [email protected] (C. Castro), [email protected] (P. MirandaAzpiazu), [email protected] (D. Fraguas), [email protected] (C. Arango). 0022-3956/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.jpsychires.2011.10.004

oxidative equilibrium is essential to neuronal differentiation and cerebral development (Fantel and Person, 2002; Do et al., 2009; Hayashi, 2009). In this vein, oxidative metabolism is increasingly being studied in neurodevelopmental and other psychiatric disorders (Mahadik and Scheffer, 1996; Chauhan and Chauhan, 2006; De Felice et al., 2009; Kapczinski et al., 2011). Recent evidence suggests that children with autism may have reduced antioxidant capacity and may suffer from chronic oxidative stress (James et al., 2008; Deth and Muratone, 2010). These studies have so far been conducted mainly in low functioning individuals with autism or in samples with a mixture of heterogeneous disorders within the autism spectrum. Oxidative stress-mediated damage to the cell membrane has also been considered a contributor to the physiopathology of schizophrenia and changes in oxidative status have been associated with the clinical course of the disorder and with response to antipsychotics (Yao et al., 2001; Akyol et al., 2002; Parikh et al., 2003).

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

To our knowledge, there has been no study investigating oxidative metabolism specifically in Asperger syndrome (AS), an Autism Spectrum Disorder (ASD) with no cognitive or language delay, or comparing this metabolism with other psychiatric disorders. The comparison of ASD (the hallmark of neurodevelopmental disorder) and psychotic disorders is relevant as, although nosologically distinct, there is increasing interest in studying the continuum between the two groups of disorders (Sporn et al., 2004; Rapoport et al., 2009; Tabares-Seisdedos and Rubenstein, 2009), including the possibility of a common pathophysiology (Crespi et al., 2010). Therefore, we aim to study oxidative status in two groups of adolescents, one with AS and another with first and recent-onset episodes of psychosis, follow them for one year, and compare them with healthy controls. Taking into account the distinctive clinical course of the two entities, with psychosis patients evolving in episodes and Asperger patients having a stable psychopathology, we hypothesized that i) excessive oxidative stress would be present both in AS and psychosis patients in comparison with control subjects, and ii) the disequilibrium would be stable for the Asperger group and dependent on symptomatology for psychosis patients. 2. Materials and methods 2.1. Participants Thirty-five adolescents with AS, 34 with a first episode of psychosis, and 34 healthy controls were recruited for this study. Asperger patients and controls were matched for age, sex, and social status. Due to disease characteristics, it was not possible to recruit a sufficient number of psychosis patients with a mean age of around 12, as in the other two groups; therefore, the youngest possible psychosis patients were recruited and age was used as a covariate in the analyses. Asperger patients were recruited through family associations, psychosis patients were recruited in the Adolescent Unit of Hospital Gregorio Marañón when they were seen for a first episode of psychosis, and healthy controls were recruited from the community. Within the group of psychosis 14 had a Schizophrenia Spectrum Disorder, 8 had a Bipolar Disorder and 12 Other Psychoses. The inclusion criteria for patients were age between 7 and 17 years at the time of first evaluation and presence of either positive psychotic symptoms (within a first psychotic episode) or a diagnosis of AS. Exclusion criteria included mental retardation (MR) per DSM-IV criteria (including not only an IQ below 70 but also impaired functioning), pervasive developmental disorder in the case of psychosis patients, neurological disorders, history of head trauma with loss of consciousness, and pregnancy. We selected healthy controls from publicly-funded schools with characteristics similar to those attended by patients. The inclusion criteria for controls were absence of a psychiatric disorder, assessed by means of the Schedule for Affective Disorders and Schizophrenia for School-Age Children-Present and Lifetime Version (K-SADS), and no neurological disorders, head trauma, pregnancy, or MR. The protocol and informed consent were approved by the Institutional Review Board at Hospital Gregorio Marañón of Madrid. All parents or legal guardians gave written informed consent before the study and patients agreed to participate in the study. 2.2. Study design All participants were assessed at baseline (time 1), 8e12 weeks after baseline (time 2) and 1 year later (time 3). At each visit, clinical measures were collected and a blood analysis was performed. A

395

general nutritional analysis was done at baseline. Non-enzymatic (glutathione and homocysteine) and enzymatic (superoxide dismutase, catalase, and glutathione peroxidase) antioxidants, total antioxidant status (TAOS), and a lipid peroxidation test, thiobarbituric acid reactive substances (TBARS) were done at baseline and 8e12 weeks. TAOS was also analyzed at 1 year.

2.3. Clinical measures Diagnosis of psychotic disorder was made according to Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV) criteria (after administering the K-SADS) (Soutullo, 1999). Patients were included in the AS group after experienced child psychiatrists (MP and CM) took a full developmental history from the parents, administered an observational interview to the patient, and applied the DSM-IV criteria (with the K-SADS to rule out concomitant psychiatric disorders) and the Gillberg criteria for AS (Gillberg and Gillberg, 1989). The full Autism Diagnostic Observation Schedule (ADOS) (Lord et al., 1989) was administered when needed (on 12 occasions), by a different clinician. The evaluating psychiatrists were experienced child psychiatrists trained in the KSADS and with a Research Certificate for the ADOS evaluation, and a diagnostic consensus was reached by the two evaluating psychiatrists in any dubious case. This diagnostic procedure was agreed upon, in view of the controversies regarding the diagnosis of Asperger syndrome (Ehlers et al., 1999; Szatmari, 2000; Mattila et al., 2007; Ghaziuddin, 2008). Sociodemographic characteristics (age, sex, and years of parental education) and treatment with antipsychotics and psychotic psychopathology at each visit, were recorded. Chlorpromazine equivalent doses were calculated for each treatment (Rijcken et al., 2003; Andreasen et al., 2010). Psychopathology was assessed with the Spanish adaptation of the Positive and Negative Symptom Scale (PANSS) (Kay et al., 1987; Peralta and Cuesta, 1994). The reliability of the different clinicians administering the PANSS was higher than 0.8 (within-class correlation coefficients).

3. Blood sample treatment, biochemical measures and analysis Fasting venous blood samples (9 mL) were collected into EDTA evacuated tubes. After immediate centrifugation, plasma and whole blood aliquots were transferred into cryogenic tubes and stored frozen at 80  C. Plasma Total Antioxidant Status was measured with a Total Antioxidant Status Assay Kit (Calbiochem, Merck KGaA, Darmstadt, Germany). Plasma lipid peroxidation was measured with a TBARS Assay Kit (Cayman Chemical, Ann Arbor, USA). The non-enzymatic antioxidants glutathione (GSH) and homocysteine (Hcy) were measured in erythrocyte lysates and plasma respectively. Reduced glutathione (GSH) was determined with a Bioxytech GSH-420 (OxisResearch). Hcy were determined using an enzymatic assay (Axis-Shield Diagnostics, Dundee, UK). Regarding enzymatic antioxidants, plasma SOD activity was measured with a Superoxide Dismutase Assay Kit (Cayman Chemical, Ann Arbor, USA). The three types of SOD (Cu/Zn-, Mn-, and Fe-SOD) were measured. The cellular glutathione peroxidase (GPx) activity was determined with a Bioxytech GPx-340 (OxisResearch, Portland, OR). The catalase activity (CAT) was measured with Bioxytech, Catalase 520, Oxis Research, Portland, OR. Both GPx and CAT were measured in erythrocyte lysates. Each of the kits was used according to the manufacturer’s instructions. A more detailed description of the biochemical analyses is provided online as Supplementary Material.

396

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

3.1. Statistical analysis Metabolic data are presented as means  SD. Normality was assessed by means of the KolmogoroveSmirnov test. All variables except for Hcy and antipsychotic doses were normally distributed. Nonparametric tests were used accordingly to assess these variables. Given that age was significantly higher in the psychosis group, correlation was sought between the metabolic variables and age and found to be negative in all cases. A one-way analysis of variance (ANCOVA with Tukey post-hoc analysis), with age and treatment at the time of assessment (dose in chlorpromazine equivalents) as covariates, was performed to ascertain whether there were inter-group (Asperger, psychosis, controls) differences between biochemical variables, separately for the 3 time points. Given that male/female ratio was different between the group with psychosis and the group with Asperger syndrome, the mean values of the oxidative metabolism variables in males and females in the whole sample was compared and found there were no statistical differences (Student t for baseline TAOS p ¼ 0.42; catalase p ¼ 0.937; superoxide dismutase p ¼ 0.705; glutathione peroxidase p ¼ 0.562 and glutathione p ¼ 0.839). Gender was also included as a covariable in the ANCOVA analysis of TAOS between groups and it did not make a difference in the results. TAOS and psychopathology at baseline were subsequently compared with those at the second visit using the Wilcoxon signedrank test. Due to sample reduction between time 2 and time 3, no comparison was performed between these two assessments. SPSS 15.0 was used for all analysis. Significance was set at p < 0.05.

Asperger, 1 psychosis, and 1 control) were taking multivitamins or mineral supplementation (in all cases below the WHO daily recommended doses) and were advised not to change doses during the study. 4.1. TAOS Total antioxidant capacity at baseline, time 2 and time 3 for the three groups of participants is presented in Table 2. Plasma total antioxidant capacity was reduced in the group of Asperger patients compared with controls at baseline and after 1 year follow-up (age used as covariate). Plasma total antioxidant capacity was reduced in the Asperger group compared with first psychotic episodes at all three assessments (age and dose of antipsychotics as covariates). Treatment with antipsychotics had no significant effect on TAOS at any of the assessments (F < 0.001, p ¼ 0.994; F ¼ 1.34, p ¼ 0.26; F ¼ 0.83, p ¼ 0.37 at baseline, time 2 and time 3 respectively). TAOS did not change from assessments at time 1 and 2 in the healthy control group (Z ¼ 0.02, p <¼ 0.988), the Asperger group (Z ¼ 1.892, p ¼ 0.059) or the psychosis group (Z ¼ 1.430, p ¼ 0.158). Regarding psychopathology, in the Asperger group, there was no change between visits 1 and 2 in either positive or negative symptoms (Z ¼ 0.251, p <¼ 0.809 and Z ¼ 1.484, p ¼ 0.141). In the psychosis group, there was a significant reduction in both positive and negative symptoms between baseline and the time 2 visit (Z ¼ 4.879, p <¼ 0.001 and Z ¼ 3.905, p <¼ 0.001, respectively). 4.2. Enzymatic and non-enzymatic antioxidants

4. Results Asperger and controls were matched for age, sex, and social status (Table 1). Although all participants in the study were in the 7e17 age range, patients in the psychosis group were 2.9 and 3.0 years older than the Asperger and healthy control groups, respectively. Parents of psychosis patients had studied a mean of 3.3 years less than parents of the Asperger and control groups. Table 1 also shows antipsychotic treatment and psychotic psychopathology at each visit. One psychosis patient was taking haloperidol and one Asperger patient was taking levomepromazine (in combination with risperidone). The rest of the patients were taking second-generation antipsychotics. Five participants (3

Levels and/or activities of the main enzymatic and nonenzymatic antioxidants measured are shown in Table 3. The activity of the antioxidant enzymes (catalase, glutathione peroxidase, and superoxide dismutase) was not different between the diagnostic groups. Nor was there a difference in plasma GSH or TBARS level between groups. Only baseline data for Hcy were available. The psychosis group had a much higher level of Hcy at baseline than any of the other groups. Other plasma antioxidants. Other baseline variables involved in the oxidant/antioxidant system are shown in Table 4. Both copper and ceruloplasmin were reduced in the psychosis group at baseline.

Table 1 Sociodemographic and clinical data.

Age Sex (M/F) Parental years of education Antipsychotic treatment Baseline Time 2a Time 3b Dose in Chlorpromazine equivalents Baseline Time 2a Time 3b PANSS positive Baseline Time 2a Time 3b PANSS negative Baseline Time 2a Time 3b

Asperger

Healthy controls

Psychosis

Between groups

Asperger vs controls

(n ¼ 35)

(n ¼ 34)

(n ¼ 34)

(p)

(p)

(p)

12.89  2.58 31/3 14.66  3.05 34.3% 27.3% 3.33% 46.2  108.4

12.79  2.87 33/2 14.09  3.21 e e e e

15.79  1.32 23/11 11.32  4.68 20.6% 88.2% 87.5% 338.5  227.45

<0.001 0.004 0.001 e e e e

0.99 0.67 0.81 e e e e

<0.001 0.004 0.001 0.158 <0.001 <0.001 <0.001

177.0  155.4 100.2  48.0 9.7  5.2 9.6  4.9 8.50, 6.04 18.0  5.8 16.7  8.5 13.3  9.4

e e e

293.8  225.6 345.5  258.2 27.4  6.0 11.5  6.5 11.3  6.9 22.6  8.2 14.1  8.4 14.2  7.8

e e e e e e e e

e e e e e e e e

0.076 0.003 <0.001 0.231 0.277 0.100 0.315 0.770

e e e

Results are shown as mean  SD. a Time 2: 8e12 weeks. Sample size at Time 2: Asperger N ¼ 33, Psychosis N ¼ 34. b Time 3: 1 year. Sample size at Time 3: Asperger N ¼ 24, Psychosis N ¼ 32.

Asperger vs Psychosis

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

397

Table 2 Plasma total antioxidant capacity (mM).

Baseline Time 2 Time 3

Asperger

Healthy controls

Psychosis

Between groups*

Asperger vs Controls*

Asperger vs Psychosis**

Psychosis vs Controls*

1.13  0.21 (32) 1.16  0.24 (30) 1.13  0.23 (16)

1.26  0.25 (33) 1.28  0.23 (31) 1.39  0.21 (8)

1.28  0.28 (25) 1.38  0.30 (32) 1.49  0.30 (21)

F ¼ 4.19; P ¼ 0.018 F ¼ 5.49; P ¼ 0.006 F ¼ 8.49; P ¼ 0.001

P ¼ 0.045 P ¼ 0.076 P ¼ 0.045

P ¼ 0.013 P ¼ 0.009 P ¼ 0.006

P ¼ 0.312 P ¼ 0.101 P ¼ 0.191

Results are shown as mean  SD. Sample size in parenthesis. ANCOVA, with age introduced as covariate (*) and age and dose of antipsychotic introduced as covariates (**). Commercial kits and lab procedures changed over the longitudinal period of the study. In order to consider only the samples processed and analyzed with the exact same methodology (as stated in the methods section), some blood samples had to be discarded. This explains the change in sample sizes between assessments.

5. Discussion Total antioxidant capacity was reduced in this group of AS individuals compared with psychosis patients and healthy controls. The reduced antioxidant capacity did not depend on any of the individual antioxidant variables measured. The result of a reduced global antioxidant capacity is quite robust (it is demonstrated at three points in time when compared with psychosis patients and at two of three times when compared with controls) and does not seem to be related to antipsychotic medication. TAOS did not change over time in the Asperger group, but contrary to our hypothesis, nor did it change in the psychosis group. Both direct and indirect data of increased oxidative stress have been reported in ASD patients (McGinnis, 2004). Plasma ROS and antioxidant activity have been shown to be altered in comparison with healthy controls (Sogut et al., 2003; Zoroglu et al., 2003, 2004; Chauhan et al., 2004b; James et al., 2004; Chauhan and Chauhan, 2006). On the other hand, metabolomic studies targeting enzymes of the homocysteine (Hcy) metabolism pathway, a critical pathway in the regulation of normal redox homeostasis and cellular methylation potential, show that methylation capacity and glutathione-dependent antioxidant/detoxification capacity are both decreased in children with autism compared with controls (Deth and Muratone, 2010). Similarly, genetic studies showing polymorphisms in the genes participating in this metabolic pathway confirm diminished methylation capacity in both patients with autism and their parents (James et al., 2006, 2008). Among various genetic elements implicated in autism, those involving the MET/PI3K pathway, highly vulnerable to oxidative stress, are also recently appointed as one of the promising candidates (Levitt and

Campbell, 2009). Direct indicators of oxidative stress, such as increased lipid peroxidation (Zoroglu et al., 2003; Chauhan et al., 2004a) and increased urinary excretion of oxidative products of DNA and fatty acids (Ming et al., 2005), have been found in ASD subjects. The results presented here add to the literature reporting that patients with AS also present the low detoxifying capacity found in severe cases of autism. Most studies do not specify the type of individuals with autism they include, often collapsing them all into an ASD group. Therefore, it is possible to assume that they are mainly low functioning patients, who represent around 70% of patients with autism. AS has been designated the milder end of a continuum of ASD (Lotspeich et al., 2004) and its biological bases have very rarely been studied. One study investigated the metabolites in the methionine cycle, the transsulfuration pathway, folate, vitamin B12, and some polymorphisms involved in the remethylation and elimination of Hcy, in three groups of children diagnosed with autistic disorders. No metabolic disturbances were found in the AS patients, while in the Autism Disorder and the nonspecific Pervasive Developmental Disorders groups, lower plasma methionine levels were observed (Pasca et al., 2008). However, the group of Asperger patients was very small (n ¼ 12). The present study did not find differences in the individual variables involved in oxidant/antioxidant metabolism between AS and either of the other groups. The literature regarding specific abnormalities in individual indicators within the redox system is contradictory. Plasma glutathione (James et al., 2004) has been previously shown to be reduced in ASD, and also the ratio of reduced to oxidized glutathione, an indicator of antioxidant activity (James et al., 2008, 2009). Regarding antioxidant enzyme activities,

Table 3 Antioxidant defences and oxidative parameters.

SOD (U/mL) Baseline Time 2 CAT (U/mL) Baseline Time 2 GPx (mU/mL) Baseline Time 2 Glutathione (mM Baseline Time 2 homocysteine TBARS (mM) Baseline Time 2

Asperger patients

Healthy controls

Psychotic patients

mean  SD

mean  SD

mean  SD

Between groups (F, p)

1.65  1.03 (32) 1.79  1.08 (30)

1.90  0.79 (33) 1.97  0. 73 (32)

1.50  0.63 (25) 1.90  0.85 (32)

1.06, 0.35 0.35, 0.71

2869.12  1278.58 (34) 2725.23  1349.57 (30)

3164.87  2089.44 (33) 3455.38  2202.12 (31)

2477.82  1569.37 (24) 2800.76  1494.79 (32)

0.86, 0.43 2.11, 0.13

31.77  19.79 (15) 31.50  17.07 (15)

33.14  15.19 (8) 47.66  30.21 (8)

22.51  18.12 (14) 32.04  22.74 (14)

0.94, 0.40 0.044, 0.96

21.45  11.72 (34) 25.06  11.04 (30) 7.21  2.19 (26)

21.32  10.12 (33) 28.09  14.96 (31) 7.76  5.44 (29)

21.84  10.82 (25) 29.69  13.78 (32) 22.23  11.99 (23)

0.028, 0.97 0.48, 0.62 21.44, <0.001a

13.62  7.88 (32) 15.09  6.77 (30)

13.06  6.93 (33) 13.33  6.24 (31)

13.53  8.92 (25) 12.86  7.28 (32)

0.09, 0.91 1.64, 0.20

Results are shown as mean  SD. Sample size in parenthesis. a Asperger vs Psychosis <0.001. *Healthy controls vs Psychosis p ¼ 0.028. Sample sizes in parentheses. TBARS: Thiobarbituric acid reactive substances; SOD: superoxide dismutase; CAT: catalase; GPx: glutathione peroxidase.

398

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

Table 4 Other antioxidant parameters at baseline. Asperger patients B12 (pg/ml) Vit A (mg/dl) Vit E (mg/dl) Albumin (g/dl) Uric acid (mg/dl) Transferrin (mg/dl) Fe (mg/dl) Ceruloplasmine (mg/dl) Cu (mg/dl) Zn (mg/dl)

540.1 43.94 1171 4.71 4.8 261.38 94.81 27 116.08 91.08

         

231.37 12.43 423.82 0.31 0.99 32.48 36.62 36.62 20.77 12.63

Healthy controls 575.86 41.17 1040.4 4.62 4.35 233.48 82.1 25.83 114.22 92.55

         

324.6 12.89 343.01 0.59 1.38 72.33 37.89 5.97 26.06 11.41

Psychotic patients 475.6 45.81 1080.8 4.72 4.9 257.11 91 22.88 94.39 83.23

         

232.42 11.12 360.32 0.38 1.55 46.19 50.29 5.98 26.4 25.51

Between groups p 0.475 0.443 0.387 0.68 0.226 0.1 0.461 0.051 0.007 0.133

P < A, 0.046 P < C, 0.02; P < A, 0.008

Results are shown as mean  SD. ANOVA with Bonferroni correction.

mainly reductions but also increases have been found in plasmatic and erythrocytic activities of glutathione peroxidase and superoxide dismutase (Yorbik et al., 2002; Sogut et al., 2003; Zoroglu et al., 2004; Al-Gadani et al., 2009), and also decreases or no change in catalase activity (Zoroglu et al., 2004; Al-Gadani et al., 2009). Ceruloplasmin and transferrin have also been found to be decreased in patients with autism, particularly in children with language regression (Chauhan et al., 2004a). Although nutritional deficiencies have been shown in autism (for various reasons, including intake selectivity), this has been the case mainly in severely affected individuals (Adams et al., 2011), and our sample was composed of subjects at the “milder” end of the spectrum. Therefore, a deficient nutritional profile was not expected and, in fact, was not observed. Reduced TAOS in AS has not, to our knowledge, been previously described. It may be a peripheral marker of nonspecific intermediate illness processes that may be triggered by various early-acting specific causes and lead to impaired neurodevelopment. 5.1. Redox system within the pathophysiology of ASD ASD have been thought to originate from deviant brain development with a strong genetic influence and a variety of etiological factors including multiple interacting risk factors involved in its physiopathology. Rarely, identified discrete biological underpinnings (Aitken, 1991) have been associated with the origin of some cases. In these, a single factor would suffice to lead to a condition such as autism. Occasionally, a genetic defect in the redox system has been shown to cause an autistic phenotype (Shoffner et al., 2010). Developmental regression is seen in up to 33% of children with autism in some reports (Goldberg et al., 2003). Environmental factors could act as precipitating agents in children with a vulnerable homeostatic background from a diverse of genetic and epigenetic origins. An immature brain or an organism with a reduced detoxifying ability would be susceptible to autistic regression (Rapin and Katzman, 1998; Keller and Persico, 2003; McGowan and Szyf, 2010). In fact, some authors argue that the effect of prenatal toxins, drugs, or infections that have been associated with autism (Folstein and Rosen-Sheidley, 2001) is possible only in the presence of genetic predisposition (Keller and Persico, 2003). The possibility is raised that commonly encountered environmental toxicants act synergistically, each at subtoxic doses, and, via their pro-oxidant activity, challenge the antioxidant or detoxifying capacity of the developing organism (Grandjean and Landrigan, 2006). The polygenic models that imply the concourse of multiple genes and environmental factors always take into account the importance of critical developmental periods for these factors to play an etiological role in the development of these

disorders. Vulnerable individuals (with suboptimal antioxidant capacity), at vulnerable developmental periods or chronically, could then suffer the consequences of a highly pro-oxidant environment and show deviances in neuronal proliferation, DNA stability and repair, gene expression, brain maturation and differentiation, and all the mechanisms that have been observed to be affected by an abnormal redox system. Taking into account the central role of mitochondria on oxidant/antioxidant balance (as  origin and also target of ROS such as O 2 and NO ), functional studies have led to its consideration as one of the possible etiological factors in autism (Lombard, 1998; Filipek et al., 2003; Deth et al., 2008; Weissman et al., 2008). The possible effect of environmental toxins in the pathogenesis of autism is one of the arguments used to explain a possible real increase in the incidence of autism (apart from the increased prevalence effect of changes in the concept and improvements in detection proficiency and diagnostic tools). Our results are therefore in keeping with previous findings suggesting increased vulnerability to oxidative stress in ASD individuals as possible contributors to the pathophysiology of autism. Although ASD have been considered mainly brain development disorders, affecting complex neural circuits, there is an increasing number of authors raising the possibility that ASD are systemic disorders (genetically influenced) with a cerebral impact (Herbert, 2005; James et al., 2008). In this sense, ASD would be a heterogeneous group of conditions with multiple different specific origins and physiopathological trajectories. These trajectories would converge in common intermediate mechanisms, such as a reduced antioxidant capacity and other physiological homeostasis, that might play a role in the deviant brain development that takes place in these disorders (Levitt and Campbell, 2009) Other metabolic aspects found in groups of patients with autism have been immunological abnormalities and abnormal methylation reactions, all interdependent (Anderson, 2008; James et al., 2008). 5.2. Autism spectrum disorders and schizophrenia spectrum disorders There is growing interest in the study of the boundaries between SSD and ASD (Rapoport et al., 2009). In fact, genetic-wise, the model that considers autism and schizophrenia to be conditions along a spectrum seems to better explain the relationships between the types of disorders than a model that consider them absolutely separate entities (Burbach and van der Zwaag, 2009; Crespi, 2010; Gauthier et al., 2010). The results of the present study show that the adolescents with psychosis have increased TAOS, increased homocysteinemia, and low Cu and ceruloplasmin. The literature shows elevated prooxidant substances, reduced antioxidant capacity, increased lipid

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

399

peroxidation and increased homocysteinemia in patients with schizophrenia. The oxidant/antioxidant imbalance has been found in stable and chronic schizophrenic patients (Ben Othmen et al., 2008), in first episode cases of schizophrenia (Mico et al., 2011), in treated and untreated patients (Mukerjee et al., 1996; Mahadik et al., 1998; Ranjekar et al., 2003), and has been inversely correlated with negative symptoms (Ustundag et al., 2006; Pazvantoglu et al., 2009). An increase in dopamine metabolism during psychotic episodes has been suggested as the origin of increased ROS and cellular damage (Hastings et al., 1996). Indeed, lipid peroxidation (an indicator of oxidative damage) has been associated with residual states of schizophrenia (Yao et al., 2001; Fenri et al., 2006). Hyperhomocysteinemia has been shown to induce oxidative stress and is considered another marker of oxidative stress (Weiss et al., 2002; Marlatt et al., 2008; Pazvantoglu et al., 2009); it has been previously associated with several neuropsychiatric disorders (Sachdev, 2004), including autism (Pasca et al., 2006), schizophrenia (Kale et al., 2010) and bipolar disorder (Dittmann et al., 2008) is known to impair neurogenesis (Rabaneda et al., 2008) and neuronal plasticity (Christie et al., 2009), and to promote neuronal degeneration (Mattson and Shea, 2003). In this regard it has been suggested that it could be a contributing factor to the pathogenesis of schizophrenia (Pasca et al., 2006). Taking the increased TAOS and Hcy found in this study together, it could be speculated that an initial antioxidant reaction of the organism takes place against the incipient psychotic process; Hcy being one of the main donors of cysteine for the synthesis of glutathione (Banner et al., 2002), its increase at this stage could serve to induce increased synthesis of glutathione. In the long term, exhaustion of the antioxidant system and damage induced by pro-oxidants as well as the concomitant increase in ROS induced by elevated Hcy could take place with subsequent low antioxidant capacity; as has been repeatedly shown in studies with chronic schizophrenia adult patients. This possibility should be tested in long-term follow-up studies. The lack of difference in total antioxidant status found between psychosis patients and controls may be due to the small number and the heterogeneous nature of our sample of psychosis patients (schizophrenia as well as other early-onset psychoses) compared with other studies comprising mainly schizophrenia patients within the psychotic group, which did find such differences. In the present study, the low antioxidant capacity of AS patients seems to be chronic (it is maintained throughout a year of followup) and part of the metabolic profile of the disorder. However, we have not been able to show that, in psychosis, the oxidant/antioxidant balance changes with disease status and/or treatment. In fact, we find that, in a first-episode psychosis group, at the beginning of the disease, antioxidant status is not reduced but rather, increased. We have previously shown that patients with a first episode of schizophrenia may have a deficient synthesis of glutathione (Mico et al., 2011), which we have not replicated in this study, and could be due to a type II error.

7. Conclusions

6. Limitations

Supplementary data related to this article can be found online at doi:10.1016/j.jpsychires.2011.10.004.

This study has several limitations: The sample may not be large enough to determine whether or not there were differences in individual antioxidant variables; the comparison psychosis group is heterogeneous and not big enough to split into subtypes such as schizophrenia and other psychoses. The nutritional assessment was determined by availability; the analysis of other endogenous antioxidant substances such as folic acid or vitamin C would render a more complete picture of the antioxidant capacity assessment. In addition, for some of the measurements, there were low numbers in some of the assessments.

In summary, we have found that a mild subtype of autism, Asperger syndrome (an autism subtype with mainly only social and communication incompetences), is characterized by low detoxifying capacity, with no specific abnormality involved. The study of mechanisms associated with oxidative stress, such as neuroinflammation, the study of upstream and downstream metabolic steps that form part of the pathophysiological trajectory that lead to the phenotype of autism, and the study of nutritional interventions that may reverse the abnormal metabolic profile, are direct derivatives from this study. Role of funding source Funding for the study came from the Ministry of Health (FIS PI040457.) and the Alicia Koplowitz Foundation. None of them was involved in the study design, collection, analysis or interpretation of the data, nor in writing the report or the decision to submit the paper. Dr Mara Parellada is under a partial research contract with the CIBERSAM, which had no further role in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decision to submit the paper for publication. Contributors Author M Parellada, C Moreno, C Arango and D Fraguas recruited the subjects, conducted the clinical interviews and the diagnostic procedures. M Parellada and C Moreno managed the literature searches. M Giraldez and C Bailon contributed to the processing of the blood samples, the management of the database and the analyses of the data. K Mc-Dowell, JC Leza, P Miranda-Azpiazu and C Castro performed the biochemical analyses of the samples and discussed the results. M Parellada conducted the statistical analyses. M Parellada, C Moreno, JC Leza, C Arango and D Fraguas discussed the main results. M Parellada wrote the first draft of the manuscript. All authors contributed to and have approved the final manuscript. Conflict of interest The authors declare no conflicts of interest in relation with the study, data or results presented. Acknowledgments Supported by CIBER de Salud Mental (CIBERSAM), Instituto de Salud Carlos III, Spanish Ministry of Science and Innovation, the Alicia Koplowitz Foundation and the Plan Nacional del Ministerio de CIencia e Innovación SAF2008-03879. Appendix. Supplementary data

References Adams JB, Audhya T, McDonough-Means S, Rubin RA, Quig D, Geis E, et al. Nutritional and metabolic status of children with autism vs. neurotypical children, and the association with autism severity. Nutrition & Metabolism (London) 2011;8:34. Adibhatla RM, Hatcher JF. Lipid oxidation and peroxidation in CNS health and disease: from molecular mechanisms to therapeutic opportunities. Antioxidants & Redox Signalling 2010;12:125e69. Aitken K. Examining the evidence for a common structural basis to autism. Developmental Medicine & Child Neurology 1991;33:930e4.

400

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401

Akyol O, Herken H, Uz E, Fadillioglu E, Unal S, Sogut S, et al. The indices of endogenous oxidative and antioxidative processes in plasma from schizophrenic patients. The possible role of oxidant/antioxidant imbalance. Progress in Neuropsychopharmacology & Biological Psychiatry 2002;26: 995e1005. Al-Gadani Y, El-Ansary A, Attas O, Al-Ayadhi L. Metabolic biomarkers related to oxidative stress and antioxidant status in Saudi autistic children. Clinical Biochemistry 2009;42:1032e40. Anderson GM. The potential role for emergence in autism. Autism Research 2008;1: 18e30. Andreasen NC, Pressler M, Nopoulos P, Miller D, Ho BC. Antipsychotic dose equivalents and dose-years: a standardized method for comparing exposure to different drugs. Biological Psychiatry 2010;67:255e62. Banner SJ, Fray AE, Ince PG, Steward M, Cookson MR, Shaw PJ. The expression of the glutamate re-uptake transporter excitatory amino acid transporter 1 (EAAT1) in the normal human CNS and in motor neurone disease: an immunohistochemical study. Neuroscience 2002;109:27e44. Ben Othmen L, Mechri A, Fendri C, Bost M, Chazot G, Gaha L, et al. Altered antioxidant defense system in clinically stable patients with schizophrenia and their unaffected siblings. Progress in Neuropsychopharmacology & Biological Psychiatry 2008;32:155e9. Burbach JP, van der Zwaag B. Contact in the genetics of autism and schizophrenia. Trends in Neurosciences 2009;32:69e72. Chauhan A, Chauhan V. Oxidative stress in autism. Pathophysiology 2006;13: 171e81. Chauhan A, Chauhan V, Brown WT, Cohen I. Oxidative stress in autism: increased lipid peroxidation and reduced serum levels of ceruloplasmin and transferrinethe antioxidant proteins. Life Sciences 2004a;75:2539e49. Chauhan V, Chauhan A, Cohen IL, Brown WT, Sheikh A. Alteration in aminoglycerophospholipids levels in the plasma of children with autism: a potential biochemical diagnostic marker. Life Sciences 2004b;74:1635e43. Christie LA, Riedel G, Platt B. Bi-directional alterations of LTP after acute homocysteine exposure. Behavioural Brain Research 2009;205:559e63. Crespi B, Stead P, Elliot M. Evolution in health and medicine Sackler colloquium: comparative genomics of autism and schizophrenia. Proceedings National Academy of Sciences U S A 2010;107(Suppl. 1):1736e41. Crespi BJ. Revisiting Bleuler: relationship between autism and schizophrenia. British Journal of Psychiatry 2010;196:495. author reply 495e496. De Felice C, Ciccoli L, Leoncini S, Signorini C, Rossi M, Vannuccini L, et al. Systemic oxidative stress in classic Rett syndrome. Free Radical Biology & Medicine 2009; 47:440e8. Deth R, Muratore C, Benzecry J, Power-Charnitsky VA, Waly M. How environmental and genetic factors combine to cause autism: a redox/methylation hypothesis. Neurotoxicology 2008;29:190e201. Deth RC, Muratone CR. The redox/methylation hypothesis of autism: a molecular mechanism for heavy metal-induced neurotoxicity. In: Chauhan A, Chauhan V, Brown T, editors. Autism oxidative stress, inflammation and immune abnormalities. Boca Raton FL CRC Press; 2010. Dittmann S, Seemuller F, Grunze HC, Schwarz MJ, Zach J, Fast K, et al. The impact of homocysteine levels on cognition in euthymic bipolar patients: a crosssectional study. Journal of Clinical Psychiatry 2008;69:899e906. Do KQ, Cabungcal JH, Frank A, Steullet P, Cuenod M. Redox dysregulation, neurodevelopment, and schizophrenia. Current Opinion in Neurobiology 2009;19: 220e30. Ehlers S, Gillberg C, Wing L. A screening questionnaire for Asperger syndrome and other high-functioning autism spectrum disorders in school age children. Journal of Autism and Developmental Disorders 1999;29:129e41. Fantel AG, Person RE. Involvement of mitochondria and other free radical sources in normal and abnormal fetal development. Annals of New York Academy of Sciences 2002;959:424e33. Fenri C, Mechri Khiari G, Othman A, Kerkeni A, Gaha L. Oxidative stress involvement in schizophrenia pathophysiology: a review. Encephale 2006;32:244e52. Filipek PA, Juranek J, Smith M, Mays LZ, Ramos ER, Bocian M, et al. Mitochondrial dysfunction in autistic patients with 15q inverted duplication. Annals of Neurology 2003;53:801e4. Folstein SE, Rosen-Sheidley B. Genetics of autism: complex aetiology for a heterogeneous disorder. Nature Review Genetics 2001;2:943e55. Gauthier J, Champagne N, Lafreniere RG, Xiong L, Spiegelman D, Brustein E, et al. De novo mutations in the gene encoding the synaptic scaffolding protein SHANK3 in patients ascertained for schizophrenia. Proceedings National Academy of Sciences U S A 2010;107:7863e8. Ghaziuddin M. Defining the behavioral phenotype of Asperger syndrome. Journal of Autism and Developmental Disorders 2008;38:138e42. Gillberg IC, Gillberg C. Asperger syndromeesome epidemiological considerations: a research note. Journal of Child Psychology and Psychiatry 1989;30:631e8. Goldberg WA, Osann K, Filipek PA, Laulhere T, Jarvis K, Modahl C, et al. Language and other regression: assessment and timing. Journal of Autism and Developmental Disorders 2003;33:607e16. Grandjean P, Landrigan PJ. Developmental neurotoxicity of industrial chemicals. Lancet 2006;368:2167e78. Hastings TG, Lewis DA, Zigmond MJ. Role of oxidation in the neurotoxic effects of intrastriatal dopamine injections. Proceedings National Academy of Sciences U S A 1996;93:1956e61. Hayashi M. Oxidative stress in developmental brain disorders. Neuropathology 2009;29:1e8.

Herbert MR. Large brains in autism: the challenge of pervasive abnormality. Neuroscientist 2005;11:417e40. James SJ, Cutler P, Melnyk S, Jernigan S, Janak L, Gaylor DW, et al. Metabolic biomarkers of increased oxidative stress and impaired methylation capacity in children with autism. American Journal of Clinical Nutrition 2004;80:1611e7. James SJ, Melnyk S, Fuchs G, Reid T, Jernigan S, Pavliv O, et al. Efficacy of methylcobalamin and folinic acid treatment on glutathione redox status in children with autism. American Journal of Clinical Nutrition 2009;89:425e30. James SJ, Melnyk S, Jernigan S, Cleves MA, Halsted CH, Wong DH, et al. Metabolic endophenotype and related genotypes are associated with oxidative stress in children with autism. American Journal of Medical Genetics B: Neuropsychiatric Genetics 2006;141B:947e56. James SJ, Melnyk S, Jernigan S, Hubanks A, Rose S, Gaylor DW. Abnormal transmethylation/transsulfuration metabolism and DNA hypomethylation among parents of children with autism. Journal of Autism and Developmental Disorders 2008;38:1966e75. Kale A, Naphade N, Sapkale S, Kamaraju M, Pillai A, Joshi S, et al. Reduced folic acid, vitamin B12 and docosahexaenoic acid and increased homocysteine and cortisol in never-medicated schizophrenia patients: implications for altered one-carbon metabolism. Psychiatry Research 2010;175:47e53. Kapczinski F, Dal-Pizzol F, Teixeira AL, Magalhaes PV, Kauer-Sant’Anna M, Klamt F, et al. Peripheral biomarkers and illness activity in bipolar disorder. Journal of Psychiatric Research 2011;45:156e61. Kay S, Opler L, Fishbein A. Positive and Negative Syndrome Scale (PANSS) rating manual. Social and Behavioral Sciences Documents. California: San Rafael; 1987. Keller F, Persico AM. The neurobiological context of autism. Molecular Neurobiology 2003;28:1e22. Levitt P, Campbell DB. The genetic and neurobiologic compass points toward common signaling dysfunctions in autism spectrum disorders. Journal of Clinical Investigation 2009;119:747e54. Lombard J. Autism: a mitochondrial disorder? Medical Hypotheses 1998;50: 497e500. Lord C, Rutter M, Goode S, Heemsbergen J, Jordan H, Mawhood L, et al. Autism diagnostic observation schedule: a standardized observation of communicative and social behavior. Journal of Autism and Developmental Disorders 1989;19: 185e212. Lotspeich LJ, Kwon H, Schumann CM, Fryer SL, Goodlin-Jones BL, Buonocore MH, et al. Investigation of neuroanatomical differences between autism and Asperger syndrome. Archives of General Psychiatry 2004;61:291e8. Mahadik SP, Mukherjee S, Scheffer R, Correnti EE, Mahadik JS. Elevated plasma lipid peroxides at the onset of nonaffective psychosis. Biological Psychiatry 1998;43: 674e9. Mahadik SP, Scheffer RE. Oxidative injury and potential use of antioxidants in schizophrenia. Prostaglandins, leukotrienes and essential fatty acids 1996;55: 45e54. Marlatt MW, Lucassen PJ, Perry G, Smith MA, Zhu X. Alzheimer’s disease: cerebrovascular dysfunction, oxidative stress, and advanced clinical therapies. Journal of Alzheimers Disease 2008;15:199e210. Mattila ML, Kielinen M, Jussila K, Linna SL, Bloigu R, Ebeling H, et al. An epidemiological and diagnostic study of Asperger syndrome according to four sets of diagnostic criteria. Journal of the American Academy of Child and Adolescent Psychiatry 2007;46:636e46. Mattson MP, Shea TB. Folate and homocysteine metabolism in neural plasticity and neurodegenerative disorders. Trends in Neuroscience 2003;26:137e46. McCord JM. The evolution of free radicals and oxidative stress. American Journal of Medicine 2000;108:652e9. McGinnis WR. Oxidative stress in autism. Alternative Therapies in Health and Medicine 2004;10:22e36. quiz 37, 92. McGowan PO, Szyf M. The epigenetics of social adversity in early life: implications for mental health outcomes. Neurobiology of Disease 2010;39:66e72. Mico JA, Rojas-Corrales MO, Gibert-Rahola J, Parellada M, Moreno D, Fraguas D, et al. Reduced antioxidant defense in early onset first-episode psychosis: a casecontrol study. BMC Psychiatry 2011;11:26. Ming X, Stein TP, Brimacombe M, Johnson WG, Lambert GH, Wagner GC. Increased excretion of a lipid peroxidation biomarker in autism. Prostaglandins, leukotrienes and essential fatty acids 2005;73:379e84. Mukerjee S, Mahadik SP, Scheffer R, Correnti EE, Kelkar H. Impaired antioxidant defense at the onset of psychosis. Schizophrenia Research 1996;19:19e26. Parikh V, Khan MM, Mahadik SP. Differential effects of antipsychotics on expression of antioxidant enzymes and membrane lipid peroxidation in rat brain. Journal of Psychiatric Research 2003;37:43e51. Pasca SP, Dronca E, Kaucsar T, Craciun EC, Endreffy E, Ferencz BK, et al. One carbon metabolism disturbances and the C667T MTHFR gene polymorphism in children with autism spectrum disorders. Journal of Cellular and Molecular Medicine; 2008. Pasca SP, Nemes B, Vlase L, Gagyi CE, Dronca E, Miu AC, et al. High levels of homocysteine and low serum paraoxonase 1 arylesterase activity in children with autism. Life Sciences 2006;78:2244e8. Pazvantoglu O, Selek S, Okay IT, Sengul C, Karabekiroglu K, Dilbaz N, et al. Oxidative mechanisms in schizophrenia and their relationship with illness subtype and symptom profile. Psychiatry and Clinical Neurosciences 2009;63:693e700. Peralta V, Cuesta MJ. Psychometric properties of the positive and negative syndrome scale (PANSS) in schizophrenia. Psychiatry Research 1994;53:31e40. Rabaneda LG, Carrasco M, Lopez-Toledano MA, Murillo-Carretero M, Ruiz FA, Estrada C, et al. Homocysteine inhibits proliferation of neuronal precursors in

M. Parellada et al. / Journal of Psychiatric Research 46 (2012) 394e401 the mouse adult brain by impairing the basic fibroblast growth factor signaling cascade and reducing extracellular regulated kinase 1/2-dependent cyclin E expression. Faseb Journal 2008;22:3823e35. Ranjekar PK, Hinge A, Hegde MV, Ghate M, Kale A, Sitasawad S, et al. Decreased antioxidant enzymes and membrane essential polyunsaturated fatty acids in schizophrenic and bipolar mood disorder patients. Psychiatry Research 2003; 121:109e22. Rapin I, Katzman R. Neurobiology of autism. Annals of Neurology 1998;43:7e14. Rapoport J, Chavez A, Greenstein D, Addington A, Gogtay N. Autism spectrum disorders and childhood-onset schizophrenia: clinical and biological contributions to a relation revisited. Journal of American Academy of Child and Adolescent Psychiatry 2009;48:10e8. Rijcken CA, Monster TB, Brouwers JR, de Jong-van den Berg LT. Chlorpromazine equivalents versus defined daily doses: how to compare antipsychotic drug doses? Journal of Clinical Psychopharmacology 2003;23:657e9. Sachdev P. [Homocysteine and neuropsychiatric disorders]. Revista Brasileira de Psiquiatria 2004;26:50e6. Shoffner J, Hyams L, Langley GN, Cossette S, Mylacraine L, Dale J, et al. Fever plus mitochondrial disease could be risk factors for autistic regression. Journal of Child Neurology 2010;25:429e34. Sogut S, Zoroglu SS, Ozyurt H, Yilmaz HR, Ozugurlu F, Sivasli E, et al. Changes in nitric oxide levels and antioxidant enzyme activities may have a role in the pathophysiological mechanisms involved in autism. Clin Chim Acta 2003;331: 111e7. Soutullo C. Traducción al Español de la Entrevista Diagnóstica: Kiddie-Schedule for Affective Disorders & Schizophrenia. In: Present & Lifetime Version (K-SADS-PL) (1996), vol. 2007; 1999. Sporn AL, Addington AM, Gogtay N, Ordonez AE, Gornick M, Clasen L, et al. Pervasive developmental disorder and childhood-onset schizophrenia:

401

comorbid disorder or a phenotypic variant of a very early onset illness? Biological Psychiatry 2004;55:989e94. Szatmari P. The classification of autism, Asperger’s syndrome, and pervasive developmental disorder. Canadian Journal of Psychiatry 2000;45:731e8. Tabares-Seisdedos R, Rubenstein JL. Chromosome 8p as a potential hub for developmental neuropsychiatric disorders: implications for schizophrenia, autism and cancer. Molecular Psychiatry 2009;14:563e89. Ustundag B, Atmaca M, Kirtas O, Selek S, Metin K, Tezcan E. Total antioxidant response in patients with schizophrenia. Psychiatry and Clinical Neurosciences 2006;60:458e64. Weiss N, Heydrick S, Zhang YY, Bierl C, Cap A, Loscalzo J. Cellular redox state and endothelial dysfunction in mildly hyperhomocysteinemic cystathionine betasynthase-deficient mice. Arteriosclerosis Thrombosis and Vascular Biology 2002;22:34e41. Weissman JR, Kelley RI, Bauman ML, Cohen BH, Murray KF, Mitchell RL, et al. Mitochondrial disease in autism spectrum disorder patients: a cohort analysis. PLoS One 2008;3:e3815. Yao JK, Reddy RD, van Kammen DP. Oxidative damage and schizophrenia: an overview of the evidence and its therapeutic implications. CNS Drugs 2001;15: 287e310. Yorbik O, Sayal A, Akay C, Akbiyik DI, Sohmen T. Investigation of antioxidant enzymes in children with autistic disorder. Prostaglandins, leukotrienes and essential fatty acids 2002;67:341e3. Zoroglu SS, Armutcu F, Ozen S, Gurel A, Sivasli E, Yetkin O, et al. Increased oxidative stress and altered activities of erythrocyte free radical scavenging enzymes in autism. European Archives of Psychiatry and Clinical Neuroscience 2004;254:143e7. Zoroglu SS, Yurekli M, Meram I, Sogut S, Tutkun H, Yetkin O, et al. Pathophysiological role of nitric oxide and adrenomedullin in autism. Cell Biochemistry and Function 2003;21:55e60.