Gene-environment interaction between an endocannabinoid system genetic polymorphism and cannabis use in first episode of psychosis.

Alterations of the endocannabinoid system (ECS) may play an important role in the development of schizophrenia and other psychotic disorders. Cannabis use is one of the environmental factors more repeatedly related to an increase the risk of developing a psychotic episode, while its use modifies the ECS normal function. In the present study we purposed to examine the gene by environment (GxE) interaction between 15 selected single nucleotide polymorphisms (SNPs) related to the ECS and cannabis use in a cohort of 321 patients with a first episode of psychosis (FEP) and 241 matched healthy controls. We found the fatty-acid amide hydrolase (FAAH) rs2295633 SNP genetic polymorphism was associated with a greater risk of presenting a FEP in subjects with relevant cannabis use, but not in subjects without a history of cannabis use. The probability of presenting a FEP was tenfold higher (OR: 10.69) in cannabis users who were homozygote carriers of the T allele of the FAAH rs2295633 SNP, compared to users of cannabis without this genotype. We also found that a higher a proportion of TT carriers of the FAAH rs2295633 SNP with a positive history of cannabis use was treated with high potency antipsychotic. This study has identified a GxE-environment interaction between a genetic polymorphism from the ECS and cannabis use involved in the risk of presenting a FEP. Although this preliminary data should be replicated with independent samples, our results highlight the importance of the pro-psychotic effects of exogenous cannabis use over the ECS in certain subjects.

A Pharmacovigilance Study in First Episode of Psychosis: Psychopharmacological Interventions and Safety Profiles in the PEPs Project.

BACKGROUND: The characterization of the first episode of psychosis and how it should be treated are principal issues in actual research. Realistic, naturalistic studies are necessary to represent the entire population of first episode of psychosis attended in daily practice. METHODS: Sixteen participating centers from the PEPs project recruited 335 first episode of psychosis patients, aged 7 to 35 years. This article describes and discusses the psychopharmacological interventions and safety profiles at baseline and during a 60-day pharmacovigilance period. RESULTS: The majority of first episode of psychosis patients received a second-generation antipsychotic (96.3%), orally (95%), and in adjusted doses according to the product specifications (87.2%). A total of 24% were receiving an antipsychotic polytherapy pattern at baseline, frequently associated with lower or higher doses of antipsychotics than the recommended ones. Eight patients were taking clozapine, all in monotherapy. Males received higher doses of antipsychotic (P=.043). A total of 5.2% of the patients were being treated with long-acting injectable antipsychotics; 12.2% of the patients received anticholinergic drugs, 12.2% antidepressants, and 13.7% mood stabilizers, while almost 40% received benzodiazepines; and 35.52% reported at least one adverse drug reaction during the pharmacovigilance period, more frequently associated with higher antipsychotic doses and antipsychotic polytherapy (85.2% vs 45.5%, P<.001). CONCLUSIONS: These data indicate that the overall pharmacologic prescription for treating a first episode of psychosis in Spain follows the clinical practice guideline recommendations, and, together with security issues, support future research of determinate pharmacological strategies for the treatment of early phases of psychosis, such as the role of clozapine, long-acting injectable antipsychotics, antipsychotic combination, and the use of benzodiazepines.

Increased susceptibility to apoptosis in cultured fibroblasts from antipsychotic-naïve first-episode schizophrenia patients.

Altered apoptosis has been proposed as a potential mechanism involved in the abnormal neurodevelopment and neurodegenerative processes associated with schizophrenia. The aim of this study was to investigate in primary fibroblast cultures whether antipsychotic-naïve patients with first-episode schizophrenia have greater apoptotic susceptibility than healthy controls. Cell growth, cell viability and various apoptotic hallmarks (caspase-3 activity, translocation of phosphatidylserine, chromatin condensation and gene expression of AKT1, BAX, BCL2, CASP3, GSK3B and P53) were measured in fibroblast cultures obtained from skin biopsies of patients (n = 11) and healthy controls (n = 8), both in basal conditions and after inducing apoptosis with staurosporine. Compared to controls, cultured fibroblasts from patients showed higher caspase-3 activity and lower BCL2 expression. When exposed to staurosporine, fibroblasts from patients also showed higher caspase-3 activity; a higher percentage of cells with translocated phosphatidylserine and condensed chromatin; and higher p53 expression compared to fibroblasts from controls. No differences in cell viability or cell growth were detected. These results strongly support the hypothesis that first-episode schizophrenia patients may have increased susceptibility to apoptosis, which may be involved in the onset and progression of the disease.

Functional analysis of gene expression in risperidone treated cells provide new insights in molecular mechanism and new candidate genes for pharmacogenetic studies.

Risperidone is a potent antagonist of both dopamine and serotonin receptors. However, little is known about the underlying molecular mechanism by which risperidone acts. Although a number of genetic variants have been observed to correlate with treatment response there are no definitive predictors of response. We performed a genome-wide gene expression analysis (Human Genome U219 Array Plate) of a human neuroblastoma cell line (SK-N-SH) exposed to risperidone to identify molecular mechanisms involved in the cellular response to risperidone and thus identify candidate genes for pharmacogenetic studies. Our results revealed that cellular risperidone treatment is associated with a range of gene expression changes, which are time (6-48h) and dose related (0.1-10µM). We found that functional clusters of these changes correspond to Gene Ontology categories related to neural cell development functions, and synaptic structure and functions. We also identified Canonical Pathways related to these functional categories: neurogenesis and axon guidance; synaptic vesicle; and neurotransmitter signaling (dopamine, serotonin and glutamate). Finally, we identified candidate genes for pharmacogenetic studies related to the main risperidone secondary effects: motor disorders, cardiovascular disorders and metabolic disorders. Our results suggest that risperidone treatment affects the neurogenesis and neurotransmission of neuroblastoma cells, which is in agreement with the "initiation and adaptation" model to explain the mechanism of action of psychotropic drugs.

Neurotoxic/neuroprotective activity of haloperidol, risperidone and paliperidone in neuroblastoma cells.

The neurotoxicity of antipsychotic (AP) drugs seems to be linked with neurological side effects like extrapyramidal symptoms (EPS). On the other hand, neuroprotective effects can mitigate or slow the progressive degenerative structural changes in the brain leading to improved outcome of schizophrenia. First and second-generation antipsychotics may differ in their neurotoxic and neuroprotective properties. The aim of this study was to compare the neurotoxic/neuroprotective activity of haloperidol, a first-generation antipsychotic, and risperidone, a second-generation one, with paliperidone, a relatively new second-generation antipsychotic, in SK-N-SH cells. Haloperidol, risperidone and paliperidone (10, 50, 100 µM) were administered, either alone or in combination with dopamine (100 µM), to human neuroblastoma SK-N-SH. We examined the effects of the drugs on cell viability (measured by alamarBlue®), caspase-3 activity (measured by fluorimetric assay) and cell death (by measuring the externalization of phosphatidylserine). Haloperidol significantly decreased cell viability and increased caspase-3 activity and cell death. Risperidone and paliperidone did not affect cell viability or cell death. Both second-generation APs decreased caspase-3 activity, especially paliperidone. In cells treated with dopamine in combination with antipsychotics, only paliperidone (10 µM) induced a slight improvement in cell viability. While haloperidol potentiated the dopamine-induced increase in caspase-3 activity, risperidone and paliperidone reduced this effect. The results indicate that haloperidol induces apoptosis, whereas risperidone and paliperidone may afford protection against it. Of the APs tested, paliperidone always showed the strongest neuroprotective effect. The different antipsychotic effects on survival and cell death might be related to differences in their capacity to induce EPS.

Sulforaphane protects SK-N-SH cells against antipsychotic-induced oxidative stress.

Adverse reactions to antipsychotic drugs (APs) have been attributed to oxidative stress. Sulforaphane (SF) is a potent antioxidant that protects against dopaminergic cell death. We examined the protective properties of SF against AP-induced oxidative stress in dopaminergic neuroblastoma cells. Human neuroblastoma SK-N-SH cells were treated with SF (0.5-5 µM), and 24 h later, haloperidol, risperidone or paliperidone (100 µM) was administered, either alone or in combination with dopamine (100 µM). To determine the antioxidant properties of SF, quinone oxidoreductase (NQO1) activity, glutathione S-transferase activity, and glutathione (GSH) levels were determined. Oxidative stress was measured by the increase in thiobarbituric acid reactive substances (TBARS) and in protein-bound quinones. Cell viability was also assessed. SF treatment increased GSH levels and induced NQO1 activity in SK-N-SH cells. Haloperidol was the only AP that increased TBARS when administered alone. When cells were cocultured with a drug in combination with dopamine, all three APs increased TBARS and protein-bound quinones and also induced neurotoxicity. In all the experimental conditions, 5 µM SF attenuated the accumulation of TBARS and protein-bound quinones and increased cell survival rates. Our results indicate that SF increases GSH levels and induces NQO1 activity and the removal of electrophilic quinones and radical oxygen species. Furthermore, SF could provide protective effects against AP-induced toxicity in dopaminergic cells.

[Systematic review of the Genomewide Association Studies (GWAS) in schizophrenia]. / Revisión sistemática de los Genomewide Association Studies (GWAS) en esquizofrenia.

INTRODUCTION: Heritability in schizophrenia can reach up to 80% and the risk in families is 5-10 times higher than in the general population. The large contribution of genetics in this disorder has led to a growing interest in its study. OBJECTIVES: To review the findings of genetic studies known as Genomewide Association Studies (GWAS) on schizophrenia. METHOD: Systematic search using Pubmed with the key words GWAS and (psychosis) or (schizophrenia). The following web pages have been reviewed: http://www.szgene.org/largescale.asp and www.genome.gov/gwastudies/. RESULTS: The GWAS have focused on causal biological aspects, such as the histocompatibility complex, glutamate metabolism, apoptosis and inflammatory processes, and the immune system (TNF-ß, TNFR1). Also focused in the search were the genes that modulate the appearance of secondary metabolic and cardiac effects and secondary effects in subjects with schizophrenia and on anti-psychotic treatment. In neurorecognition, over-expression of the MET proto-oncogene (MET) has been associated with a low susceptibility for schizophrenia and a better cognitive performance, as well as a lower susceptibility for the incidence of cancer. Mention is also made of the different genes that mediate in cognitive functioning depending on the anti-psychotic treatment received. CONCLUSIONS: The main interests of the GWAS during the last few years have been the neurobiological pathways involved in schizophrenia. The discoveries arising from these studies have been limited. This has led to an innovative approach on the aetiological study of the disorder by studying gene-environment interactions.

Hnf1alpha (MODY3) controls tissue-specific transcriptional programs and exerts opposed effects on cell growth in pancreatic islets and liver.

Heterozygous HNF1A mutations cause pancreatic-islet beta-cell dysfunction and monogenic diabetes (MODY3). Hnf1alpha is known to regulate numerous hepatic genes, yet knowledge of its function in pancreatic islets is more limited. We now show that Hnf1a deficiency in mice leads to highly tissue-specific changes in the expression of genes involved in key functions of both islets and liver. To gain insights into the mechanisms of tissue-specific Hnf1alpha regulation, we integrated expression studies of Hnf1a-deficient mice with identification of direct Hnf1alpha targets. We demonstrate that Hnf1alpha can bind in a tissue-selective manner to genes that are expressed only in liver or islets. We also show that Hnf1alpha is essential only for the transcription of a minor fraction of its direct-target genes. Even among genes that were expressed in both liver and islets, the subset of targets showing functional dependence on Hnf1alpha was highly tissue specific. This was partly explained by the compensatory occupancy by the paralog Hnf1beta at selected genes in Hnf1a-deficient liver. In keeping with these findings, the biological consequences of Hnf1a deficiency were markedly different in islets and liver. Notably, Hnf1a deficiency led to impaired large-T-antigen-induced growth and oncogenesis in beta cells yet enhanced proliferation in hepatocytes. Collectively, these findings show that Hnf1alpha governs broad, highly tissue-specific genetic programs in pancreatic islets and liver and reveal key consequences of Hnf1a deficiency relevant to the pathophysiology of monogenic diabetes.

-141C Ins/Del polymorphism of the dopamine D2 receptor gene is associated with schizophrenia in a Spanish population.

OBJECTIVE: In this study we examined the relationship between dopamine D2 receptor (DRD2) polymorphisms (TaqIA, TaqIB, -141C Ins/Del) and dopamine D3 receptor (DRD3) Ser9Gly polymorphism and the risk of schizophrenia in a Spanish population. METHODS: Two hundred and forty-three schizophrenia patients and 291 healthy controls from the general population participated in a case-control study. RESULTS: No significant differences were observed in the allele or genotype frequencies of TaqIA, TaqIB or Ser9Gly polymorphisms between the schizophrenia patients and the healthy controls. The frequency of the -141C Del allele was significantly lower in the former group (odds ratio=0.4, P=0.01). The -141C Del allele, which produces lower expression of DRD2, may protect against dopaminergic hyperactivity in schizophrenia. CONCLUSION: This study is one of the few studies of Caucasian participants that supports the results obtained in the original Japanese study, in which the -141C Ins/Del polymorphism was first described. Furthermore, our findings reinforce the hypothesis that excess dopaminergic activity leads to schizophrenia.

Induction of apoptosis in HT-29 cells by extracts from isothiocyanates-rich varieties of Brassica oleracea.

Among the vegetables with anti-carcinogenic properties, members of the genus Brassica are the most effective at reducing the risk of cancer. This property may be explained by their principle bioactive compounds, isothiocyanates (ITCs). The aim of this study was to measure the amounts of ITCs in extracts from vegetables of the Brasssica genus and assay them for potency of induction of apoptosis in a colorectal cancer cell line (HT-29). ITCs were determined by the cyclocondensation assay with 1,2-benzenedithiol and induction of apoptosis by assessment of cell viability, caspase-3 activity and DNA fragmentation. Purple cabbage extract showed the highest ITC concentration per gram, fresh weight, followed by black cabbage and Romanesco cauliflower. At ITC concentrations of 7.08 microg/mL these extracts decreased cell viability and induced caspase-3 and DNA fragmentation at 48h. Brussels sprouts showed the strongest effects on cell viability and caspase-3 activity. Varieties of Brassica Oleracea are rich sources of ITCs that potently inhibit the growth of colon cancer cells by inducting apoptosis. All the extracts showed anticancer activity at ITC concentrations of between 3.54 to 7.08 mug/mL, which are achievable in vivo. Our results showed that ITC concentration and the chemopreventive responses of plant extracts vary among the varieties of Brassica Oleracea studied and among their cultivars.

Lower specific micronutrient intake in colorectal cancer patients with tumors presenting promoter hypermethylation in p16(INK4a), p4(ARF) and hMLH1.

BACKGROUND: The diversity of the Mediterranean diet and the heterogeneity of acquired epigenetic alterations in colorectal cancer (CRC) led us to examine the possible association between dietary factors and promoter hypermethylation in genes implicated in the pathogenesis of these neoplasms (p16(INK4a), p14(ARF), hMLH1) and the interaction with methylene tetrahydrofolate reductase (MTHFR) C677T polymorphism. PATIENTS AND METHODS: For the molecular study, 120 CRC patients were analyzed for hMLH1 promoter methylation status and MTHFR genotyping. Dietary patterns and molecular data on p16(INK4a) and p14(ARF) methylation were obtained from previous studies with this populations. RESULTS: Patients with methylation in p16(INK4a) consumed significantly less folate (p = 0.01), vitamin A (p = 0.01), vitamin B1 (p = 0.007), potassium (p = 0.03) and iron (p = 0.02) than controls. Patients with methylation in p14(ARF) or hMLH1 consumed significantly less vitamin A (p = 0.001 and p = 0.05, respectively). CONCLUSION: These results support that certain micronutrients protect against colorectal neoplasia and emphasize the importance of considering the different molecular forms of CRC as etiologically distinct diseases.

Induction of NAD(P)H quinone oxidoreductase by vegetables widely consumed in Catalonia, Spain.

Monofunctional inducers (MIs) enhance phase 2 enzymes such as nicotinamide-adenine-dinucleotide-phosphate [NAD(P)H] quinone oxidoreductase (NQO1) without modifying oxidation enzymes. The induction of these protective enzymes appears to be mediated by genetic regulatory elements in their promoter regions known as the antioxidant response element (ARE). The aim of this study was to identify, through an in vitro study, which of the 30 fruits and vegetables commonly consumed in Catalonia, Spain, contain MIs of NQO1. We assayed the capacity of extracts of these fruits and vegetables to induce NQO1 [by more than 1.5-fold: ratio of induction (cells treated/control) >1.5, 8-mg/ml dose] in two murine hepatoma cell lines: Hepa 1c1c7 and BPrC1, a modified cell line that possesses a nonfunctional aryl hydrocarbon receptor nuclear translocator system and is thus nonresponsive to bifunctional inducers. We also used a third cell line, papiloma (PE) murine keratinocytes, a stably transfected cell line with an ARE-luc+ plasmid (AREPE cell line) for verifying induction through the ARE with a simple luminescence screening assay. Broccoli (Hepa 1c1c7, ratio=5.5; BPrC1, ratio=2.3), calcot (Allium cepa L.) (Hepa 1c1c7, ratio=4.7; BPrC1, ratio=.5), green onion (Hepa 1c1c7, ratio=4.6; BPrC1, ratio=2), green cabbage (Hepa 1c1c7, ratio=3.6; BPrC1, ratio=2.7), purple cabbage (Hepa 1c1c7, ratio=3.4; BPrC1, ratio=2), and black cabbage (Hepa 1c1c7, ratio=3; BPrC1, ratio=3) were active NQO1 inducers in both murine hepatoma cell lines. Extracts from broccoli (ratio=3.5), calcot (ratio=4.8), cauliflower (ratio=4.2), cabbage (ratio=2.2), green onion (ratio=3.2), green cabbage (ratio=3.6), black cabbage (ratio=4.5), and purple cabbage (ratio=3.7) were confirmed to contain MIs in the AREPE cell line. These results are very similar to those described for vegetables consumed in the United States, with the exception of calcot, which is common in Catalonia but is not grown or consumed widely in the United States.

Decrease in specific micronutrient intake in colorectal cancer patients with tumors presenting Ki-ras mutation.

BACKGROUND: The diversity of the Mediterranean diet and the heterogeneity of acquired genetic alterations in colorectal cancer (CRC) led us to examine the possible association between dietary factors and mutations, such as Ki-ras mutations, in genes implicated in the pathogenesis of these neoplasms. PATIENTS AND METHODS: The study was based on 246 cases and 296 controls. For the molecular study only 117 patients with Ki-ras tumor expression were included. Dietary patterns were assessed using a semi-quantitative food frequency questionnaire. RESULTS: Patients with Ki-ras mutations in codon 12 (K12) consumed significantly less vitamin A (p=0.02), B1 (p=0.01), D (p=0.02) and iron (p=0.03) than controls, whereas patients without these mutations had similar intakes of these nutrients to controls. The consumption of fiber, folate, vitamin E and potassium was lower in the two subgroups of patients (K12-positive or -negative) than in controls. Mutation in codon 13 was not associated with any nutrient deficit. CONCLUSION: These results support previous findings that certain micronutrients protect against colorectal neoplasia and emphasize the importance of considering the different molecular forms of CRC as etiologically distinct diseases.

High-resolution analysis of DNA copy number alterations in colorectal cancer by array-based comparative genomic hybridization.

Array-based comparative genomic hybridization (CGH) allows for the simultaneous examination of thousands of genomic loci at 1-2 Mb resolution. Copy number alterations detected by array-based CGH can aid in the identification and localization of cancer causing genes. Here we report the results of array-based CGH in a set of 125 primary colorectal tumors hybridized onto an array consisting of 2463 bacterial artificial chromosome clones. On average, 17.3% of the entire genome was altered in our samples (8.5 +/- 6.7% gained and 8.8 +/- 7.3% lost). Losses involving 8p, 17p, 18p or 18q occurred in 37, 46, 49 and 60% of cases, respectively. Gains involving 8q or 20q were observed 42 and 65% of the time, respectively. A transition from loss to gain occurred on chromosome 8 between 41 and 48 Mb, with 25% of cases demonstrating a gain of 8p11 (45-53 Mb). Chromosome 8 also contained four distinct loci demonstrating high-level amplifications, centering at 44.9, 60, 92.7 and 144.7 Mb. On 20q multiple high-level amplifications were observed, centering at 32.3, 37.8, 45.4, 54.7, 59.4 and 65 Mb. Few differences in DNA copy number alterations were associated with tumor stage, location, age and sex of the patient. Microsatellite stable and unstable (MSI-H) tumors differed significantly with respect to the frequency of alterations (20 versus 5%, respectively, P < 0.01). Interestingly, MSI-H tumors were also observed to have DNA copy number alterations, most commonly involving 8q. This high-resolution analysis of DNA copy number alterations in colorectal cancer by array-based CGH allowed for the identification of many small, previously uncharacterized, genomic regions, such as on chromosomes 8 and 20. Array-based CGH was also able to identify DNA copy number changes in MSI-H tumors.

Pooled analysis and meta-analysis of glutathione S-transferase M1 and bladder cancer: a HuGE review.

Smoking is a known risk factor for bladder cancer. The product of the GSTM1 gene, glutathione S-transferase M1 (GSTM1), is involved in the detoxification of polycyclic aromatic hydrocarbons found in tobacco smoke; a homozygous deletion of this gene in approximately 50% of Caucasians and Asians results in a lack of GSTM1 enzyme activity. Most studies examining the relation between bladder cancer and GSTM1 have reported an increased risk associated with a lack of GSTM1 activity. The authors performed meta- and pooled analyses of published and unpublished, case-control, genotype-based studies that examined this association (17 studies, 2,149 cases, 3,646 controls) and excluded studies conducted in populations with a high prevalence of exposure to known bladder cancer risk factors other than tobacco smoke. Using random effects models in the meta-analysis, the authors obtained a summary odds ratio of 1.44 (95% confidence interval (CI): 1.23, 1.68) for GSTM1 null status with all studies included. Results from studies with at least 100 cases and 100 controls produced a summary odds ratio of 1.42 (95% CI: 1.26, 1.60). Pooled analyses using original data sets from 10 studies (1,496 cases and 1,444 controls) and adjusting for age, sex, and race produced similar results. There was no evidence of multiplicative interaction between the GSTM1 null genotype and ever smoking in relation to bladder cancer, although there was a suggestion of additive interaction (additive interaction = 0.45, 95% CI: -0.03, 0.93). These results indicate that, among populations studied to date, GSTM1 null status is associated with a modest increase in the risk of bladder cancer.