Low expression of catecholamine-O-methyl-transferase gene in obsessive-compulsive disorder

Journal of Anxiety Disorders 23 (2009) 660–664

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Journal of Anxiety Disorders

Low expression of catecholamine-O-methyl-transferase gene in obsessive-compulsive disorder Zhen Wang a,b,c,1,*, Zeping Xiao a, Sabra S. Inslicht b,c, Huiqi Tong c, Wenhui Jiang a, Xiao Wang a, Thomas Metzler c, Charles R. Marmar b,c, Sanduo Jiang a a b c

Shanghai Mental Health Center, Shanghai Jiao Tong University, China Department of Psychiatry, University of California, San Francisco, USA Veterans Affairs Medical Center, San Francisco, USA

A R T I C L E I N F O

A B S T R A C T

Article history: Received 20 March 2008 Received in revised form 27 January 2009 Accepted 4 February 2009

This study examined peripheral catecholamine-O-methyl-transferase (COMT) gene expression in obsessive-compulsive disorder (OCD) patients and healthy controls. Participants included 35 first episode OCD patients and 31 age- and sex-matched healthy controls. Relative COMT gene expression levels were examined by real-time quantitative reverse transcription polymerase chain reaction (RTPCR) in peripheral blood of all the subjects. COMT gene expression levels, normalized by glyceraldehyde3-phosphate dehydrogenase (GAPDH), were significantly decreased in the OCD group compared with healthy controls (F = 6.244, p = 0.015). OCD patients showed a 32% down-regulation. We also found lower COMT gene expression levels in female in comparison to male participants (F = 5.366, p = 0.024) in the sample as a whole. COMT gene expression down-regulation of male OCD patients relative to male controls is 38%, and that of female OCD patients relative to female controls is 27%. These results suggest that COMT gene expression down-regulation might play an important role in the development of OCD and that there may be gender differences in this alteration. ß 2009 Elsevier Ltd. All rights reserved.

Keywords: Catecholamine-O-methyl-transferase Obsessive-compulsive disorder Gene expression RT-PCR

1. Introduction Obsessive-compulsive disorder (OCD) is a common, severe, and chronically debilitating mental illness that affects 1–3% of the general population (Eisen et al., 1999; Weissman et al., 1994). This disorder is characterized by recurrent obsessions and/or compulsions that are severe enough to be time-consuming or cause marked distress or significant impairment in functioning (American Psychiatric Association, 2000). Although etiology of this disorder remains largely unknown, numerous family (Black, Noyes, Goldstein, & Blum, 1992; Nicolini, Weissbecker, Mejia, & Sanchez de Carmona, 1993; Pauls et al., 1995) and twin studies (Andrews, Stewart, Allen, & Henderson, 1990; Rasmussen & Tsuang, 1986; Rosario-Campos et al., 2001) suggest that OCD has a strong genetic component. The candidate gene approach is a common method used by investigators to explore the genetic etiology of OCD. Based on the

* Corresponding author at: Shanghai Mental Health Center, 600 Wan Ping Nan Road, Shanghai 200030, China. Tel.: +86 21 64387250; fax: +86 21 64387986. E-mail address: [email protected] (Z. Wang). 1 Current address: Veterans Affairs Medical Center, 4150 Clement Street (116P), San Francisco, CA 94121, USA. Tel.: +1 415 221 4810x3531; fax: +1 415 751 2297. 0887-6185/$ – see front matter ß 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.janxdis.2009.02.004

evidence that dopamine antagonists are beneficial for treatmentrefractory OCD patients (Denys, van Megen, & Westenberg, 2002; McDougle, Epperson, Pelton, Wasylink, & Price, 2000; Ramasubbu, Ravindran, & Lapierre, 2000) and agents enhancing dopamine release may induce obsessive-compulsive symptoms (Crum & Anthony, 1993; Satel & McDougle, 1991), the catecholamine-Omethyl-transferase (COMT) gene has been suggested as one of the most important candidate genes. COMT is an enzyme that has a crucial role in the regulation of both the dopaminergic and noradrenergic neurotransmitter systems. COMT is encoded by a single gene localized on chromosome 22q11 region, which has a close relationship with a variety of mental disorders including OCD (Carlson et al., 1997; Gothelf et al., 2004, 2007; Murphy, 2002). The COMT gene has a common functional polymorphism at codon 158 where a nucleotide transition (G to A) causes a change in the amino acid sequence (Val to Met) in the protein product. Multiple studies assessing the relationship between this polymorphism and OCD yield contrasting results. Some have found that this polymorphism was associated with OCD in female (Alsobrook et al., 2002) or male patients (Denys, Van Nieuwerburgh, Deforce, & Westenberg, 2006; Karayiorgou et al., 1997; Pooley, Fineberg, & Harrison, 2007); yet, several studies and one meta-analysis found no association of this gene polymorphism in OCD (Azzam & Mathews, 2003; Erdal et al., 2003; Meira-Lima et al., 2004). While

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other functional variations in this gene have been associated with mental diseases, the results are inconsistent and those variations have not been examined in an OCD population (Dempster, Mill, Craig, & Collier, 2006; Li et al., 2000; Sanders et al., 2005). It is still not clear whether this important gene contains specific functional variants mediating vulnerability to OCD. Recent research has demonstrated that complex ‘epigenetic’ factors, which regulate gene function without altering the DNA code, may a key role in the development of mental disease (Tsankova, Renthal, Kumar, & Nestler, 2007). These may explain inconsistent results found in association studies and also imply that gene function, such as gene expression, may play a more important role in the etiology of mental disorders. However, prior studies on the relation of OCD and COMT gene have only focused on gene polymorphisms rather than on gene function. In this pilot study, we examine the association between OCD and COMT gene expression using realtime quantitative reverse transcription polymerase chain reaction (RT-PCR). 2. Methods and materials

661

Table 1 Characteristics of patients and healthy controls. OCD

Controls

t/x2

p

Sex Male Female

19 16

17 14

0.02

1.00

Age

31.2  12.0

30.3  5.8

0.38

0.70

Marital status Married Unmarried Divorce

13 20 2

15 16 0

2.35

0.31

Age of onset

26.9  11.9

NA

Y-BOCS Obsession (1–5 items) Compulsion (6–10 items)

21.5  5.4 11.2  5.0 10.3  6.1

NA NA NA

Chatsworth, CA, USA). Reverse transcription was performed using TaqMan reverse transcription reagents with random primers (Takara Bio, Kyoto, Japan). Samples were stored at 70 8C prior to further use.

2.1. Subjects 2.3. Real-time quantitative RT-PCR Participants included 35 patients with OCD and 31 age- and sex-matched healthy controls. All subjects were Han nationality Chinese and had no consanguinity. OCD patients were recruited from the outpatient mental health clinic of the Shanghai Mental Health Center (SMHC). We used the SMHC clinical interview which is a semi-structured intake tool used widely throughout the hospital to evaluate the presence of DSM-IV axis I disorders and additional Chinese-specific diagnoses. Diagnostic interviews were conducted by two attending psychiatrists, with one serving as the primary interviewer and the other making independent ratings simultaneously during the interview. OCD patients were included for participation if they met first episode OCD, had a minimum Yale-Brown Obsessive-Compulsive Scale (Y-BOCS; Kim, Dysken, & Kuskowski, 1990) score of 16, did not meet any other DSM-IV axis I diagnosis and did not receive previous pharmacotherapy or psychotherapy for OCD symptoms. Forty-two patients were referred from SMHC clinic and 35 of them met all of the inclusion criteria and agreed to participate in this study. Seven patients were excluded because they had a prior diagnosis of OCD (n = 3), were receiving psychotherapy (n = 1), have received pharmacotherapy (n = 1) or met criteria for panic disorder (n = 1) or alcohol abuse (n = 1). Healthy controls included staff members of the Shanghai Mental Health Center and medical students of Fudan University and Shanghai Jiao Tong University. All the controls were interviewed by a psychiatrist to exclude any DSM-IV axis I diagnosis (the interview procedure was the same as that of the OCD group except only one interviewer was present). Participants (OCD patients and healthy cases) were included if they were at least 18 years of age, were without serious somatic disease, and had completed at least a middle school education. There were no differences in gender, age, marital status, as shown in Table 1. This study was approved by the Ethics Committee of Shanghai Mental Health Center and written informed consent was obtained from all subjects. 2.2. Sample collection Blood samples were obtained from all subjects for extraction of mRNA. Total RNA was extracted from peripheral whole blood using the QIAamp RNA blood Mini Kit (QIAGEN, Chatsworth, CA, USA) following the manufacturer’s standard protocol. To ensure no DNA contamination, clean-up of the RNA was performed using QIAGEN spin columns with an additional DNAase step (QIAGEN,

Real-time quantitative RT-PCR was performed in triplicate for each sample on an ABI Prism 7900sequence detection system with TaqMan Universal PCR mastermix (Applied Biosystems, Foster City, CA, USA) according to the manufacturer’s protocol. An AssayBy-Demand probe/primer set specific to COMT (Hs00241349_m1) and housekeeping control gene (glyceraldehyde-3-phosphate dehydrogenase, GAPDH) probe were purchased from Applied Biosystems (Applied Biosystems, Foster City, CA, USA). The expression data produced were analyzed and converted into threshold cycle values (Ct-values) using SDS 2.0 (Applied Biosystems, Foster City, CA, USA). Ct value was defined as the cycle number at which the fluorescence exceeds the preset fluorescence threshold. The average Ct-value of GAPDH and COMT was calculated from triplicate results for each sample. The comparative Ct method (DCt) was used to measure the relative gene expression (Johnson, Wang, Smith, Heslin, & Diasio, 2000). The comparative Ct method is a quantitation approach to assess relative changes in mRNA levels between two samples. The DCtvalue of each sample (patients and controls) was obtained by subtracting the average GAPDH Ct-value of each sample from the average COMT Ct-value of each sample (i.e., COMT expression normalized by GAPDH). The gene expression level was calculated using the equation 2DC t (Cui, Jiang, Jiang, Xu, & Yao, 2005; Zhu et al., 2004). Subsequently, the average DCt of healthy control subjects was used as calibrator and the fold change (how many times more the gene is expressed in one group than in another group) was calculated by the formula 2DDC t (DDCt was DCt of calibrator minus the DCt of individual patients). 2.4. Statistical analysis Statistical analyses were conducted using SPSS for Windows (Version 13.0). Two-way ANOVAs were used to compare the COMT expression level between groups and genders, and to test for an interaction between group and gender. T-Tests were performed to compare groups on continuous variables and Chi-square tests were used for categorical variables. 3. Results The effects of group and gender on COMT gene expression levels were analyzed using a two-way ANOVA (Table 2). There was a

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Table 2 Two-way analysis of variance for group and gender effects on COMT gene expression level. Source

Df

SS

MS

F

P

Group Gender Group  gender Error

1 1 1 62

0.002 0.002 0.001 0.023

0.002 0.002 0.001 0.000

6.244 5.366 2.291

0.015 0.024 0.135

Total

66

0.132

Table 3 Fold change of COMT gene expression. Calibrators (relative expression = 1)

Target groups

Fold changes

Controls (N = 31) Female controls (N = 14) Male controls (N = 17)

OCD (N = 35) Female OCD (N = 16) Male OCD (N = 19)

1.32 1.27 1.38

significant difference between the two groups (F = 6.244, p = 0.015) and between genders (F = 5.366, p = 0.024), but no group by gender interaction (F = 2.291, p = 0.135). COMT gene expression levels, normalized by GAPDH, were significantly decreased in the OCD group compared with healthy control subjects. COMT gene expression level was lower for female subjects in comparison to male subjects. If we divided the subjects into four subgroups according to group and gender, the COMT gene expression level was ordered as following: male controls (N = 17, 0.045  0.022), female controls (N = 14, 0.036  0.009), male OCD (N = 19, 0.035  0.015), and female OCD (N = 16, 0.032  0.011). We have not further compared the gene expression differences among these four groups because of sample size limitations. DD The 2 C (fold changes) of individual OCD patients were calculated by taking the average DCt of healthy control subjects as a calibrator. On comparing with healthy controls (relative expression = 1), the COMT expression fold change of OCD patients was 1.32, indicating a 32% COMT expression down-regulation. Taking the average DCt of female controls as a calibrator for female OCD patients and the average DCt of male controls as a calibrator for male OCD patients, revealed that the average COMT expression fold change of male OCD patients was 1.38 (or 38% down regulation) and 1.27 (or 28% down regulation) in female OCD patients (Table 3). There were no significant Pearson correlations between COMT expression level and OCD severity on the Y-BOCS (r = 0.09, p = 0.62). 4. Discussion This is the first study to date that examines gene expression of COMT in peripheral blood of OCD patients. We found that COMT gene expression level, normalized by GAPDH, was significantly decreased in OCD patients when compared with healthy controls. Using the average DCt of age and gender matched healthy control subjects as a calibrator, OCD patients were 32% lower on COMT expression level. COMT is involved in the inactivation of the catecholamine neurotransmitters dopamine, epinephrine and norepinephrine, by introducing a methyl group to these catecholamines which is donated by S-adenosyl methionine (SAM). COMT is very important for regulating prefrontal cortex (PFC) function (Goldman-Rakic, Muly, & Williams, 2000) and cognitive processing (Joober et al., 2002) by modulating dopamine levels in the PFC (Tunbridge, Harrison, & Weinberger, 2006). This is particularly significant for OCD since dopaminergic function and PFC function are thought to be dysregulated in this disorder (Friedlander & Desrocher, 2006; Maiho¨fner et al., 2007; Russell et al., 2003; van der Wee et al., 2004),

and changes of COMT activity and function are hypothesized to be mechanisms involved in OCD. Since COMT gene expression level is a relatively effective index to reflect COMT function, we hypothesized that COMT gene expression would be altered in patients with OCD. The present study results revealed gene expression alterations of COMT in OCD patients, which may directly affect dopamine metabolism and cause concentration changes in the cortex. The low level of COMT mRNA expression in OCD patients reflects the decrease of total COMT activity and may increase dopamine signaling in brain. There is substantial evidence suggesting that OCD symptoms are related to increased dopamine neurotransmission (Denys, Zohar, & Westenberg, 2004). Our findings suggest that dopamine dysfunction is involved in the pathogenesis of OCD, supporting the dopamine hypothesis of the pathophysiology of OCD (Westenberg, Fineberg, & Denys, 2007). Most existing polymorphism studies of COMT and OCD report a gender difference of COMT val158 met polymorphism. Four case control studies reported an association between the met158 allele of COMT and OCD in men but not in women while only one study found association with OCD in women. In this study, we also found a significant gender difference in COMT gene expression, COMT gene expression level o female subjects was lower than male. In this study, we did not find a significant correlation between COMT expression level and OCD severity. This may be due to the relative homogeneity of this patient group with respect to COMT expression since strict inclusion criteria were implemented. It is also possible that COMT expression level may result from a combination of several factors which we did not measure in this study, such as psychological stress and cognitive functioning. For OCD patients, the relevant functional change of COMT and dopamine occur in the brain. We measured COMT peripherally in blood, since obtaining tissue from the relevant brain regions is essentially impossible for reasonably large and representative patient sample sets. Can the gene expression of peripheral blood, a more accessible tissue, reflect gene expression in the CNS? Middleton et al. (2005) first conducted a study on schizophrenia and bipolar disorder using quantitative real-time RT-PCR, and found compelling evidence for the utility of analyzing peripheral blood leukocytes RNA as a marker for changes in expression in neuropsychiatric disorders. A recent study (Sullivan, Fan, & Perou, 2006) used gene expression data from 79 diverse human tissues for 33,698 gene transcripts to address the broad question of whether peripheral blood lymphocyte gene expression data is a representative and reasonable surrogate for gene expression in the CNS. The results suggested that gene expression in lymphocytes is relatively strongly correlated with gene expression in brain. A study of human COMT expression in brain and lymphoblast also showed a strong correlation of COMT expression between brain and blood (Zhu et al., 2004). These results indicate that peripheral blood markers reflect relevant expression and functional changes of COMT in the brain. Numerous candidate genes of OCD, such as COMT, 5-HTT, DRD4, 5-HT2A, have been studied over the past two decades (Hemmings & Stein, 2006), but have not achieved breakthrough results until now. A growing number of studies suggest that gene function may play a more important role in the etiology of neuropsychiatric disorders (Hemmings & Stein, 2006; Tsankova et al., 2007). To our knowledge only one semi-quantitative gene expression study with a small sample size has been carried out in OCD (Rocca et al., 2000). Our results suggest that gene expression of OCD is a promising marker for the disorder and should be further studied with larger samples in the future. While our results are promising, there are several limitations that should be considered. First, we only performed one method, real-time reverse transcription PCR, to assess COMT gene expression. Although it is often described as a ‘‘gold’’ standard,

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future studies research should consider examining regulatory RNAs, protein levels and protein activity (Nolan, Hands, & Bustin, 2006). Second, OCD was not diagnosed with standardized measures although DSM-IV criteria were implemented. Third, the control group was selected from a highly educated population, which may have been different from the patient sample. Although gender and age are matched, we cannot exclude the possibility that other confounds may have impacted our results such as life stresses and IQ. Fourth, although there is compelling evidence that gene expression in blood can be strongly correlated with gene expression in the brain, it is still an indirect approach to approximate brain function. In conclusion, we have examined the role of COMT gene expression in OCD for the first time. We have demonstrated lower COMT expression in peripheral blood in OCD patients and gender differences in COMT expression. Future studies are needed to replicate the current findings and to follow-up the relationships between COMT expression and treatment outcome. Conflict of interest None. Acknowledgements This research was supported by the grants from Shanghai Health Bureau (044Y26) and Shanghai Clinical Medicine Center of Psychiatry, PR China (K-04-3). The authors thank all the participants in this study, Dr. Carol Mathews and Thomas C. Neylan for valuable suggestion on the manuscript and Ms. Y.P. Qian for help with data analysis and laboratory work. References Alsobrook, J. P., II, Zohar, A. H., Leboyer, M., Chabane, N., Ebstein, R., & Pauls, D. L. (2002). Association between the COMT locus and obsessive-compulsive disorder in females but not males. American Journal of Medical Genetics, 114, 116–120. American Psychiatric Association. (2000). Obsessive compulsive disorder Diagnostic and statistical manual of mental disorders, (4th edition-text revision). Washington, DC: APA Press. p. 456–463. Andrews, G., Stewart, G., Allen, R., & Henderson, A. S. (1990). The genetics of six neurotic disorders: a twin study. Journal of Affective Disorders, 19, 23–29. Azzam, A., & Mathews, C. A. (2003). Meta-analysis of the association between the catecholamine-O-methyl-transferase gene and obsessive-compulsive disorder. American Journal of Medical Genetics B (Neuropsychiatric Genetic), 123, 64–69. Black, D. W., Noyes, R., Jr., Goldstein, R. B., & Blum, N. (1992). A family study of obsessive-compulsive disorder. Archives of General Psychiatry, 49, 362–368. Carlson, C., Papolos, D., Pandita, R. K., Faedda, G. L., Veit, S., Goldberg, R., et al. (1997). Molecular analysis of velo-cardio-facial syndrome patients with psychiatric disorders. American Journal of Human Genetics, 60, 851–859. Crum, R. M., & Anthony, J. C. (1993). Cocaine use and other suspected risk factors for obsessive-compulsive disorder: a prospective study with data from the Epidemiologic Catchment Area surveys. Drug and Alcohol Dependence, 31, 281–295. Cui, D. H., Jiang, K. D., Jiang, S. D., Xu, Y. F., & Yao, H. (2005). The tumor suppressor adenomatouspolyposis coli gene is associated with susceptibility to schizophrenia. Molecular Psychiatry, 10, 669–677. Dempster, E. L., Mill, J., Craig, I. W., & Collier, D. A. (2006). The quantification of COMT mRNA in post mortem cerebellum tissue: diagnosis, genotype, methylation and expression. BMC Medical Genetics, 7, 10. Denys, D., van Megen, H., & Westenberg, H. (2002). Quetiapine addition to serotonin reuptake inhibitor treatment in patients with treatment-refractory obsessivecompulsive disorder: an open-label study. Journal of Clinical Psychiatry, 63, 700– 703. Denys, D., Van Nieuwerburgh, F., Deforce, D., & Westenberg, H. (2006). Association between the dopamine D2 receptor TaqI A2 allele and low activity COMT allele with obsessive-compulsive disorder in males. European Neuropsychopharmacology, 16, 446–450. Denys, D., Zohar, J., & Westenberg, H. G. (2004). The role of dopamine in obsessivecompulsive disorder: preclinical and clinical evidence. Journal of Clinical Psychiatry, 65(Suppl 14), 11–17. Eisen, J. L., Goodman, W. K., Keller, M. B., Warshaw, M. G., DeMarco, L. M., Luce, D. D., et al. (1999). Patterns of remission and relapse in obsessive compulsive disorder: a 2-year prospective study. Journal of Clinical Psychiatry, 60, 346–351. Erdal, M. E., Tot, S., Yazici, K., Yazici, A., Herken, H., Erdem, P., et al. (2003). Lack of association of catechol-O-methyltransferase gene polymorphism in obsessivecompulsive disorder. Depression and Anxiety, 18, 41–45.

663

Friedlander, L., & Desrocher, M. (2006). Neuroimaging studies of obsessive-compulsive disorder in adults and children. Clinical Psychology Review, 26, 32–49. Goldman-Rakic, P. S., Muly, E. C., 3rd, & Williams, G. V. (2000). D(1) receptors in prefrontal cells and circuits. Brain Research Brain Research Reviews, 31, 295–301. Gothelf, D., Michaelovsky, E., Frisch, A., Zohar, A. H., Presburger, G., Burg, M., et al. (2007). Association of the low-activity COMT 158 Met allele with ADHD and OCD in subjects with velocardiofacial syndrome. International Journal of Neuropsychopharmacology, 10, 301–308. Gothelf, D., Presburger, G., Zohar, A. H., Burg, M., Nahmani, A., Frydman, M., et al. (2004). Obsessive-compulsive disorder in patients with velocardiofacial (22q11 deletion) syndrome. American Journal of Medical Genetics B (Neuropsychiatric Genetic), 126, 99–105. Hemmings, S. M., & Stein, D. J. (2006). The current status of association studies in obsessive-compulsive disorder. Psychiatric Clinics of North America, 29, 411– 444. Johnson, M. R., Wang, K., Smith, J. B., Heslin, M. J., & Diasio, R. B. (2000). Quantitation of dihydropyrimidinedehydrogenase expression by real-time reverse transcription polymerase chain reaction. Analytical Biochemistry, 278, 175–184. Joober, R., Gauthier, J., Lal, S., Bloom, D., Lalonde, P., Rouleau, G., et al. (2002). Catechol-O-methyltransferase Val-108/158-Met gene variants associated with performance on the Wisconsin Card Sorting Test. Archives of General Psychiatry, 59, 662–663. Karayiorgou, M., Altemus, M., Galke, B. L., Goldman, D., Murphy, D. L., Ott, J., et al. (1997). Genotype determining low catechol-O-methyltransferase activity as a risk factor for obsessive-compulsive disorder. Proceedings of the National Academy of Sciences of the United States of America, 94, 4572–4575. Kim, S. W., Dysken, M. W., & Kuskowski, M. (1990). The Yale-Brown obsessivecompulsive scale: a reliability and validity study. Psychiatry Research, 34, 99– 106. Li, T., Ball, D., Zhao, J., Murray, R. M., Liu, X., Sham, P. C., et al. (2000). Family-based linkage disequilibrium mapping using SNP marker haplotypes: application to a potential locus for schizophrenia at chromosome 22q11. Molecular Psychiatry, 5, 77–84. Maiho¨fner, C., Sperling, W., Kaltenha¨user, M., Bleich, S., de Zwaan, M., Wiltfang, J., et al. (2007). Spontaneous magnetoencephalographic activity in patients with obsessive-compulsive disorder. Brain Research, 1129, 200–205. McDougle, C. J., Epperson, C. N., Pelton, G. H., Wasylink, S., & Price, L. H. (2000). A double-blind, placebo-controlled study of risperidone addition in serotonin reuptake inhibitor-refractory obsessive-compulsive disorder. Archives of General Psychiatry, 57, 794–801. Meira-Lima, I., Shavitt, R. G., Miguita, K., Ikenaga, E., Miguel, E. C., & Vallada, H. (2004). Association analysis of the catechol-o-methyltransferase (COMT), serotonin transporter (5-HTT) and serotonin 2A receptor (5HT2A) gene polymorphisms with obsessive-compulsive disorder. Genes, Brain and Behavior, 3, 75–79. Middleton, F. A., Pato, C. N., Gentile, K. L., McGann, L., Brown, A. M., Trauzzi, M., et al. (2005). Gene expression analysis of peripheral blood leukocytes from discordant sib-pairs with schizophrenia and bipolar disorder reveals points of convergence between genetic and functional genomic approaches. American Journal of Medical Genetics B (Neuropsychiatric Genetic), 136, 12–25. Murphy, K. C. (2002). Schizophrenia and velo-cardio-facial syndrome. Lancet, 359, 426– 430. Nicolini, H., Weissbecker, K., Mejia, J. M., & Sanchez de Carmona, M. (1993). Family study of obsessive-compulsive disorder in a Mexican population. Archives of Medical Research, 24, 193–198. Nolan, T., Hands, R. E., & Bustin, S. A. (2006). Quantification of mRNA using real-time RT-PCR. Nature Protocols, 1, 1559–1582. Pauls, D. L., Alsobrook, J. P., Phil, M., Goodman, W., Rasmussen, S., & Leckman, J. F. (1995). A family study of obsessive-compulsive disorder. American Journal of Psychiatry, 152, 76–84. Pooley, E. C., Fineberg, N., & Harrison, P. J. (2007). The met(158) allele of catechol-Omethyltransferase (COMT) is associated with obsessive-compulsive disorder in men: case-control study and meta-analysis. Molecular Psychiatry, 12, 556–561. Ramasubbu, R., Ravindran, A., & Lapierre, Y. (2000). Serotonin and dopamine antagonism in obsessive-compulsive disorder: effect of atypical antipsychotic drugs. Pharmacopsychiatry, 33, 236–238. Rasmussen, S. A., & Tsuang, M. T. (1986). Clinical characteristics and family history in DSM-III obsessive compulsive disorder. American Journal of Psychiatry, 143, 317– 322. Rocca, P., Beoni, A. M., Eva, C., Ferrero, P., Maina, G., Bogetto, F., et al. (2000). Lymphocyte peripheral benzodiazepine receptor mRNA decreases in obsessivecompulsive disorder. European Neuropsychopharmacology, 10, 337–340. Rosario-Campos, M. C., Leckman, J. F., Mercadante, M. T., Shavitt, R. G., Prado, H. S., Sada, P., et al. (2001). Adults with early-onset obsessive-compulsive disorder. American Journal of Psychiatry, 158, 1899–1903. Russell, A., Cortese, B., Lorch, E., Ivey, J., Banerjee, S. P., Moore, G. J., et al. (2003). Localized functional neurochemical marker abnormalities in dorsolateral prefrontal cortex in pediatric obsessive-compulsive disorder. Journal of Child and Adolescent Psychopharmacology, 13(Suppl 1), 31–38. Sanders, A. R., Rusu, I., Duan, J., Vander Molen, J. E., Hou, C., Schwab, S. G., et al. (2005). Haplotypic association spanning the 22q11.21 genes COMT and ARVCF with schizophrenia. Molecular Psychiatry, 10, 353–365. Satel, S. L., & McDougle, C. J. (1991). Obsessions and compulsions associated with cocaine abuse [Letter to the Editor]. American Journal of Psychiatry, 148, 947. Sullivan, P. F., Fan, C., & Perou, C. M. (2006). Evaluating the comparability of gene expression in blood and brain. American Journal of Medical Genetics (Neuropsychiatric Genetic), 141, 261–268.

664

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Tsankova, N., Renthal, W., Kumar, A., & Nestler, E. J. (2007). Epigenetic regulation in psychiatric disorders. Nature Reviews Neuroscience, 8, 355–367. Tunbridge, E. M., Harrison, P. J., & Weinberger, D. R. (2006). Catechol-o-methyltransferase, cognition, and psychosis: Val158Met and beyond. Biological Psychiatry, 60, 141–151. van der Wee, N. J., Stevens, H., Hardeman, J. A., Mandl, R. C., Denys, D. A., van Megen, H. J., et al. (2004). Enhanced dopamine transporter density in psychotropic-naive patients with obsessive-compulsive disorder shown by (123I){beta}-CIT SPECT. American Journal of Psychiatry, 161, 2201–2206.

Weissman, M. M., Bland, R. C., Canino, G. J., Greenwald, S., Hwu, H. G., Lee, C. K., et al. (1994). The cross national epidemiology of obsessive-compulsive disorder. Journal of Clinical Psychiatry, 53(Suppl 3), 5–10. Westenberg, H. G., Fineberg, N. A., & Denys, D. (2007). Neurobiology of obsessivecompulsive disorder: serotonin and beyond. CNS Spectrums, 12(2 Suppl 3), 14– 27. Zhu, G., Lipsky, R. H., Xu, K., Ali, S., Hyde, T., Kleinman, J., et al. (2004). Differential expression of human COMT alleles in brain and lymphoblasts detected by RTcoupled 50 nuclease assay. Psychopharmacology, 177, 178–184.