Depression, Cytokine, and Cytokine by Treatment Interactions Modulate Gene Expression in Antipsychotic Naïve First Episode Psychosis.

In schizophrenia, genetic and environmental factors affect neurodevelopment and neuroprogressive trajectory. Altered expression of neuro-immune genes and increased levels of cytokines are observed, especially in patients with comorbid depression. However, it remains unclear whether circulating levels of cytokines and expression of these genes are associated, and how antipsychotic treatments impact this association. Relationships between messenger RNA (mRNA) expression of 11 schizophrenia-related genes and circulating levels of cytokines (interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α) were analyzed in 174 antipsychotic naïve first episode psychosis (FEP) and in 77 healthy controls. A subgroup of 72 patients was reassessed after treatment with risperidone. FEP patients were divided into those with and without depression. FEP patients with depression showed increased COMT expression and decreased NDEL1 expression. Increased IL-6 was associated with lowered AKT1 and DROSHA expression, while increased IL-10 was associated with increased NDEL1, DISC1, and MBP expression. IL-6 levels significantly increased the risperidone-induced expression of AKT1, DICER1, DROSHA, and COMT mRNA. The differential mRNA gene expression in FEP is largely associated with increased cytokine levels. While increased IL-6 may downregulate AKT-mediated cellular functions and dysregulate genes involved in microRNA (miRNA) machinery, increased IL-10 has neuroprotective properties. Increased IL-6 levels may prime the expression of genes (AKT1, DICER1, DROSHA, and COMT) in response to risperidone, suggesting that cytokine × treatment × gene interactions may improve cell function profiles. FEP patients with depression show a different gene expression profile reinforcing the theory that depression in FEP is a different phenotype.

Effect of APOE and CHRNA7 genotypes on the cognitive response to cholinesterase inhibitor treatment at different stages of Alzheimer's disease.

The loss of cholinergic transmission is considered to be an important cause of Alzheimer's disease (AD). Treatment with acetyl cholinesterase inhibitors (ChEIs) shows benefits; however, great heterogeneity has been observed in patient responses. We evaluated apolipoprotein E (APOE) and α7 nicotinic receptor (CHRNA7) single-nucleotide polymorphisms (SNPs) and associated these SNPs with pharmacological responses to ChEIs in a Brazilian population with AD. We studied 177 outpatients using ChEIs, and they were classified as responders and nonresponders according to variation in Mini-Mental State Examination (MMSE) status. The analysis of APOE genotypes showed that patients with the ε4 allele had a worse response than those without the ε4 allele. We observed an association between the CHRNA7 T allele and a better response to treatment with ChEIs in patients with mild AD (MMSE ≥ 20). The SNP rs6494223 of CHRNA7 as well as APOEε4 could be useful for understanding the response to ChEI treatment in patients with AD.

Changes in gene expression and methylation in the blood of patients with first-episode psychosis.

Schizophrenia is a severe mental health disorder with high heritability. The investigation of individuals during their first-episode psychosis (FEP), before the progression of psychotic disorders and especially before treatment with antipsychotic medications, is particularly helpful for understanding this complex disease and for the identification of potential biomarkers. In this study, we compared the expression of genes that are involved in neurotransmission and neurodevelopment of antipsychotic-naive FEP in the peripheral blood of patients (n=51) and healthy controls (n=51). In addition, we investigated the differentially expressed genes with respect to a) DNA methylation, b) the correlation between gene expression and clinical variables (PANSS), and c) gene expression changes after risperidone treatment. Expression levels of 11 genes were quantified with SYBR Green. For methylation analysis, bisulfite sequencing was performed. A significant decrease in GCH1 mRNA levels was observed in FEP patients relative to controls. Also, when we compare the FEP patients after risperidone treatment with controls, this difference remains significant, and no significant differences were observed in GCH1 mRNA levels when comparing patients before and after risperidone treatment. Additionally, although the differences were non-significant after Bonferroni correction, the expression of GCH1 seemed to be correlated with PANSS scores, and the GCH1 promoter region was more methylated in FEP than in controls, thus corroborating the results obtained at the mRNA level. Few studies have been conducted on GCH1, and future studies are needed to clarify its potential role in the progression of schizophrenia.

Expression profile of neurotransmitter receptor and regulatory genes in the prefrontal cortex of spontaneously hypertensive rats: relevance to neuropsychiatric disorders.

The spontaneously hypertensive rat (SHR) strain was shown to be a useful animal model to study several behavioral, pathophysiological and pharmacological aspects of schizophrenia and attention-deficit/hyperactivity disorder. To further understand the genetic underpinnings of this model, our primary goal in this study was to compare the gene expression profile of neurotransmitter receptors and regulators in the prefrontal cortex (PFC) and nucleus accumbens (NAcc) of SHR and Wistar rats (control group). In addition, we investigated DNA methylation pattern of promoter region of the genes differentially expressed. We performed gene expression analysis using a PCRarray technology, which simultaneously measures the expression of 84 genes related to neurotransmission. Four genes were significantly downregulated in the PFC of SHR compared to Wistar rats (Gad2, Chrnb4, Slc5a7, and Qrfpr) and none in nucleus accumbens. Gad2 and Qrfpr have CpG islands in their promoter region. For both, the promoter region was hypomethylated in SHR group, and probably this mechanism is not related with the downregulation of these genes. In summary, we identified genes that are downregulated in the PFC of SHR, and might be related to the behavioral abnormalities exhibited by this strain.

Effect of antipsychotic drugs on gene expression in the prefrontal cortex and nucleus accumbens in the spontaneously hypertensive rat (SHR).

Antipsychotic drugs (APDs) are the standard treatment for schizophrenia. The therapeutic effect of these drugs is dependent upon the dopaminergic D2 blockade, but they also modulate other neurotransmitter pathways. The exact mechanisms underlying the clinical response to APDs are not fully understood. In this study, we compared three groups of animals for the expression of 84 neurotransmitter genes in the prefrontal cortex (PFC) and nucleus accumbens (NAcc). Each group was treated with a different APD (risperidone, clozapine or haloperidol), and with a non-treated group of spontaneously hypertensive rats (SHRs), which is an animal model for schizophrenia. This study also explored whether or not differential expression was regulated by DNA methylation in the promoter region (PR). In the clozapine group, we found that Chrng was downregulated in the NAcc and six genes were downregulated in the PFC. In the haloperidol group, Brs3 and Glra1 were downregulated, as was Drd2 in the clozapine group and Drd3, Galr3 and Gabrr1 in the clozapine and haloperidol groups. We also encountered four hypermethylated CG sites in the Glra1 PR, as well as three in the risperidone group and another in the haloperidol group, when compared to non-treated rats. Following the APD treatment, the gene expression results revealed the involvement of genes that had not previously been described, in addition to the activity of established genes. The investigation of the involvement of these novel genes can lead to better understanding about the specific mechanisms of action of the individual APDs studied.

Evaluation of neurotransmitter receptor gene expression identifies GABA receptor changes: a follow-up study in antipsychotic-naïve patients with first-episode psychosis.

A study of the gene expression levels in the blood of individuals with schizophrenia in the beginning of the disease, such as first-episode psychosis (FEP), is useful to detect gene expression changes in this disorder in response to treatment. Although a large number of genetic studies on schizophrenia have been conducted, little is known about the effects of antipsychotic treatment on gene expression. The aim of the present study was to examine differences in the gene expression in the blood of antipsychotic-naïve FEP patients before and after risperidone treatment (N = 44) and also to verify the correlation with treatment response. In addition, we determined the correlations between differentially expressed genes and clinical variables. The expression of 40 neurotransmitter and neurodevelopment-associated genes was assessed using the RT2 Profiler PCR Array. The results indicated that the GABRR2 gene was downregulated after risperidone treatment, but no genes were associated with response to treatment and clinical variables after Bonferroni correction. GABRR2 downregulation after treatment can both suggest an effect of risperidone treatment or processes related to disease progression, either not necessarily associated with the improvement of symptoms. Despite this change was observed in blood, this decrease in GABRR2 mRNA levels might be an effect of changes in GABA concentrations or other systems interplay consequently to D2 blockage induced by risperidone, for example. Thus, it is important to consider that antipsychotics or the progression of psychotic disorders might interfere with gene expression.

Analysis of HSPA8 and HSPA9 mRNA expression and promoter methylation in the brain and blood of Alzheimer's disease patients.

Alzheimer's disease (AD) is the most common form of dementia in elderly. Chaperones may have a crucial role in AD due to their involvement in protein quality control, folding, and degradation. In this study, we investigated the mRNA and promoter DNA methylation levels of two chaperones, HSPA8 and HSPA9, in postmortem brain tissue (entorhinal and auditory cortices and hippocampus) from healthy elderly and AD subjects as well as in peripheral blood of healthy elderly and AD patients. mRNA quantification was performed by qRT-PCR and DNA methylation by mass spectrometry. In the peripheral blood, we did not observe a significant difference in HSPA8 and HSPA9 expression between elderly controls and AD. A significant downregulation of HSPA8 and HSPA9 was observed in AD across the three brain regions compared to the controls, suggesting their participation in AD pathogenesis. However, no important DNA methylation differences were observed, suggesting that other mechanism may be involved in controlling these genes expression.

Neurotransmitter receptor and regulatory gene expression in peripheral blood of Brazilian drug-naïve first-episode psychosis patients before and after antipsychotic treatment.

Little is known about how genes expressed in blood relate to schizophrenia or antipsychotic use. We analyzed gene expression in 10 first-episode psychosis patients and nine controls using PCR Arrays. GABRR2 and CHRNA3 were found to be differentially expressed after risperidone treatment. These genes may be regulated by antipsychotic use.

CNP and DPYSL2 mRNA expression and promoter methylation levels in brain of Alzheimer's disease patients.

Alzheimer's disease (AD) is a highly prevalent type of dementia in the elderly population. AD is a complex neurodegenerative disorder. Thus, epigenetic mechanisms that regulate gene expression might have an important role in AD. CNP (2',3'-Cyclic Nucleotide 3' Phosphodiesterase) gene encodes a protein used as an index of myelin alterations. DPYSL2 (Dihydropyrimidinase-like 2) is described as acting in structural and regulatory processes in the central nervous system, such as neural differentiation, neurotransmitter release, and stabilization of microtubules. In this study, we evaluated gene expression and epigenetic regulation of CNP and DPYSL2 genes in three postmortem brain regions (entorhinal and auditory cortices and hippocampus) of AD patients and healthy elderly controls. mRNA quantification was performed using qRT-PCR, and promoter DNA methylation patterns were determined by mass spectrometry using the Sequenom EpiTYPER platform. We observed CNP mRNA downregulation in entorhinal and auditory cortex in relation to the same regions of the control group. CNP alterations in the brain might suggest impairment in myelination leading to a synaptic and cognition loss. No AD-associated differences in CNP and DPYSL2 promoter DNA methylation were observed, suggesting that other mechanisms may be involved in mediating the observed CNP gene expression.

SORL1 and SIRT1 mRNA expression and promoter methylation levels in aging and Alzheimer's Disease.

Alzheimer's Disease (AD) is a neurodegenerative disorder and the most common cause of dementia among the elderly. Efforts have been made to understand the genetic and epigenetic mechanisms involved in the development of this disease. As SORL1 (sortilin-related receptor) and SIRT1 (sirtuin 1) genes have been linked to AD pathogenesis, we aimed to investigate their mRNA expression and promoter DNA methylation in post mortem brain tissues (entorhinal and auditory cortices and hippocampus) from healthy elderly subjects and AD patients. We also evaluated these levels in peripheral blood leukocytes from young, healthy elderly and AD patients, investigating whether there was an effect of age on these profiles. The comparative CT method by Real Time PCR and MALDI-TOF mass spectrometry were used to analyze gene expression and DNA methylation, respectively. SORL1 gene was differently expressed in the peripheral blood leukocytes and might act as a marker of aging in this tissue. Furthermore, we found that SORL1 promoter DNA methylation might act as one of the mechanisms responsible for the differences in expression observed between blood and brain for both healthy elderly and AD patients groups. The impact of these studied genes on AD pathogenesis remains to be better clarified.

SMARCA5 methylation and expression in gastric cancer.

Here, we first evaluated SMARCA5 expression and promoter DNA methylation in gastric carcinogenesis. Immunohistochemistry and methylation-specific PCR were analyzed in 19 and 48 normal mucosa and in 52 and 92 gastric cancer samples, respectively. We observed higher immunoreactivity of SMARCA5 in gastric cancer samples than in normal mucosa. Moreover, SMARCA5 promoter methylation was associated with the absence of protein expression. Our findings suggest that SMARCA5 has a potential role in proliferation and malignancy in gastric carcinogenesis.

Insulin-like growth factor binding protein-3 gene methylation and protein expression in gastric adenocarcinoma.

OBJECTIVE: The aim of this study was to evaluate IGFBP-3 protein expression, its correlation with gene promoter methylation pattern in gastric carcinogenesis and with clinicopathological characteristics. DESIGN: Forty-three normal gastric mucosa and 94 adenocarcinoma samples were investigated through methylation specific PCR, after bisulfite modification. Immunohistochemistry was analyzed using peroxidase in 54 gastric cancer and 20 normal gastric mucosa samples. RESULTS: IGFBP-3 expression was higher in tumor samples than in normal mucosa (p<0.0001). Intestinal type presented a higher frequency of protein expression than diffuse type (p=0.0412). Methylation frequency of IGFBP-3 promoter in gastric samples revealed, respectively, 95.7% and 97.7% in neoplastic and non-neoplastic samples. The frequency of IGFBP-3 methylation did not differ between tumor and normal samples (95.7% versus 97.7%, p=0.7810). We did not observe a significant correlation between IGFBP-3 promoter methylation and protein expression. CONCLUSION: In summary, our study did not observe any influence of IGFBP-3 promoter methylation on protein expression. Moreover we propose that IGFBP-3 immunostaining in gastric tissue may be a useful marker for malignancy.

hTERT methylation and expression in gastric cancer.

Gastric cancer is the second most prevalent cause of cancer death worldwide. DNA methylation is a common event in gastric carcinogenesis. hTERT seems to be the rate-limiting determinant of telomerase activation, which is responsible for stability and life span. hTERT hypermethylation has been associated with telomerase expression. In the present study, we investigated the promoter methylation status and hTERT protein expression in gastric cancer and normal mucosa samples. One hundred and nine gastric cancer and 53 normal mucosa samples were investigated through methylation-specific PCR. Immunohistochemistry was analysed using peroxidase in 55 gastric cancer and 18 normal gastric mucosa samples. This is the first study evaluating hTERT methylation status in gastric carcinogenesis. We did not observe hTERT protein expression in normal gastric mucosa. Moreover, hTERT expression was observed in 80% of tumours and was associated with gastric cancer (p < 0.0001). Partial methylation was the most frequent pattern in gastric samples, even in normal mucosa. The frequency of specimens presenting hypermethylation was significantly higher in tumours than in normal mucosa samples (p = 0.0002), although the presence of hypermethylated promoter was not associated with a higher frequency of hTERT expression. A low correlation between hTERT protein expression and methylation was verified in gastric cancer samples. There was a clear difference in the frequency of hTERT expression and methylation within tumoral and non-tumoral tissues. Methylation status and telomerase expression may be useful for the diagnosis of gastric cancer and may have an impact on the anti-telomerase strategy for cancer therapy.

Promoter methylation analysis of SIRT3, SMARCA5, HTERT and CDH1 genes in aging and Alzheimer's disease.

Longevity related genes were investigated concerning promoter methylation. SIRT3, SMARCA5, HTERT and CDH1 promoters were analyzed in peripheral blood in relation to gender, age and Alzheimer's disease (AD). Methylation Specific PCR assay (MSP) was used. There were no significant differences in methylation frequencies of SIRT3, SMARCA5 and CDH1 among young, elderly and AD groups (p> 0.05), showing no association with aging or AD. On the other hand, HTERT methylation frequency was associated with the aging process, in that AD patients differed from elderly controls (p=0.0086), probably due to telomere and immune dysfunctions involved in AD pathogenesis.

Promoter hypermethylation of CDH1, FHIT, MTAP and PLAGL1 in gastric adenocarcinoma in individuals from Northern Brazil.

AIM: To evaluate the methylation status of CDH1, FHIT, MTAP and PLAGL1 promoters and the association of these findings with clinico-pathological characteristics. METHODS: Methylation-specific PCR (MSP) assay was performed in 13 nonneoplastic gastric adenocarcinoma, 30 intestinal-type gastric adenocarcinoma and 35 diffuse-type gastric adenocarcinoma samples from individuals in Northern Brazil. Statistical analyses were performed using the chi-square or Fisher's exact test to assess associations between methylation status and clinico-pathological characteristics. RESULTS: Hypermethylation frequencies of CDH1, FHIT, MTAP and PLAGL1 promoter were 98.7%, 53.9%, 23.1% and 29.5%, respectively. Hypermethylation of three or four genes revealed a significant association with diffuse-type gastric cancer compared with nonneoplastic cancer. A higher hypermethylation frequency was significantly associated with H pylori infection in gastric cancers, especially with diffuse-type. Cancer samples without lymph node metastasis showed a higher FHIT hypermethylation frequency. MTAP hypermethylation was associated with H pylori in gastric cancer samples, as well as with diffuse-type compared with intestinal-type. In diffuse-type, MTAP hypermethylation was associated with female gender. CONCLUSION: Our findings show differential gene methylation in tumoral tissue, which allows us to conclude that hypermethylation is associated with gastric carcinogenesis. MTAP promoter hypermethylation can be characterized as a marker of diffuse-type gastric cancer, especially in women and may help in diagnosis, prognosis and therapies. The H pylori infectious agent was present in 44.9% of the samples. This infection may be correlated with the carcinogenic process through the gene promoter hypermethylation, especially the MTAP promoter in diffuse-type. A higher H pylori infection in diffuse-type may be due to greater genetic predisposition.