P2RX7: Expression Responds to Sleep Deprivation and Associates with Rapid Cycling in Bipolar Disorder Type 1

Context

Rapid cycling is a severe form of bipolar disorder with an increased rate of episodes that is particularly treatment-responsive to chronotherapy and stable sleep-wake cycles. We hypothesized that the P2RX7 gene would be affected by sleep deprivation and be implicated in rapid cycling.

Objectives

To assess whether P2RX7 expression is affected by total sleep deprivation and if variation in P2RX7 is associated with rapid cycling in bipolar patients.

Design

Gene expression analysis in peripheral blood mononuclear cells (PBMCs) from healthy volunteers and case-case and case-control SNP/haplotype association analyses in patients.

Participants

Healthy volunteers at the sleep research center, University of California, Irvine Medical Center (UCIMC), USA (n = 8) and Swedish outpatients recruited from specialized psychiatric clinics for bipolar disorder, diagnosed with bipolar disorder type 1 (n = 569; rapid cycling: n = 121) and anonymous blood donor controls (n = 1,044).

Results

P2RX7 RNA levels were significantly increased during sleep deprivation in PBMCs from healthy volunteers (p = 2.3*10⁻⁹). The P2RX7 rs2230912 _A allele was more common (OR = 2.2, p = 0.002) and the ACGTTT haplotype in P2RX7 (rs1718119 to rs1621388) containing the protective rs2230912_G allele (OR = 0.45–0.49, p = 0.003–0.005) was less common, among rapid cycling cases compared to non-rapid cycling bipolar patients and blood donor controls.

Conclusions

Sleep deprivation increased P2RX7 expression in healthy persons and the putatively low-activity P2RX7 rs2230912 allele A variant was associated with rapid cycling in bipolar disorder. This supports earlier findings of P2RX7 associations to affective disorder and is in agreement with that particularly rapid cycling patients have a more vulnerable diurnal system.     

Marked reduction of AKT1 expression and deregulation of AKT1-associated pathways in peripheral blood mononuclear cells of schizophrenia patients

Background

Recent studies have suggested that deregulated AKT1 signaling is associated with schizophrenia. We hypothesized that if this is indeed the case, we should observe both decreased AKT1 expression as well as deregulation of AKT1 regulated pathways in Peripheral Blood Mononuclear Cells (PBMCs) of schizophrenia patients.

Objectives

To examine PBMC expression levels of AKT1 in schizophrenia patients versus controls, and to examine whether functional biological processes in which AKT1 plays an important role are deregulated in schizophrenia patients.

Methods/Results

A case-control study, investigating whole-genome PBMC gene expression in male, recent onset (<5 years) schizophrenia patients (N = 41) as compared to controls (N = 29). Genes, differentially expressed between patients and controls were identified using ANOVA with Benjamini-Hochberg correction (false discovery rate (FDR) = 0.05). Functional aspects of the deregulated set of genes were investigated with the Ingenuity Pathway Analysis (IPA) Software Tool. We found significantly decreased PBMC expression of AKT1 (p<0.001, t = −4.25) in the patients. AKT1 expression was decreased in antipsychotic-free or -naive patients (N = 11), in florid psychotic (N = 20) and in remitted (N = 21) patients. A total of 1224 genes were differentially expressed between patients and controls (FDR = 0.05). Functional analysis of the entire deregulated gene set indicated deregulated canonical pathways involved in a large number of cellular processes: immune system, cell adhesion and neuronal guidance, neurotrophins and (neural) growth factors, oxidative stress and glucose metabolism, and apoptosis and cell-cycle regulation. Many of these processes are associated with AKT1.

Conclusions

We show significantly decreased PBMC gene expression of AKT1 in male, recent-onset schizophrenia patients. Our observations suggest that decreased PBMC AKT1 expression is a stable trait in recent onset, male schizophrenia patients. We identified several AKT related cellular processes which are potentially affected in these patients, a majority of which play a prominent role in current schizophrenia hypotheses.     

Hepatic expression patterns of inflammatory and immune response genes associated with obesity and NASH in morbidly obese patients

Background

Obesity modulates inflammation and activation of immune pathways which can lead to liver complications. We aimed at identifying expression patterns of inflammatory and immune response genes specifically associated with obesity and NASH in the liver of morbidly obese patients.

Methodology/Principal Findings

Expression of 222 genes was evaluated by quantitative RT-PCR in the liver of morbidly obese patients with histologically normal liver (n = 6), or with severe steatosis without (n = 6) or with NASH (n = 6), and in lean controls (n = 5). Hepatic expression of 58 out of 222 inflammatory and immune response genes was upregulated in NASH patients. The most notable changes occurred in genes encoding chemokines and chemokine receptors involved in leukocyte recruitment, CD and cytokines involved in the T cell activation towards a Th1 phenotype, and immune semaphorins. This regulation seems to be specific for the liver since visceral adipose tissue expression and serum levels of MCP1, IP10, TNFα and IL6 were not modified. Importantly, 47 other genes were already upregulated in histologically normal liver (e.g. CRP, Toll-like receptor (TLR) pathway). Interestingly, serum palmitate, known to activate the TLR pathway, was increased with steatosis.

Conclusion/Significance

The liver of obese patients without histological abnormalities already displayed a low-grade inflammation and could be more responsive to activators of the TLR pathway. NASH was then characterized by a specific gene signature. These findings help to identify new potential actors of the pathogenesis of NAFLD.     

Gene expression and biological pathways in tissue of men with prostate cancer in a randomized clinical trial of lycopene and fish oil supplementation

Background

Studies suggest that micronutrients may modify the risk or delay progression of prostate cancer; however, the molecular mechanisms involved are poorly understood. We examined the effects of lycopene and fish oil on prostate gene expression in a double-blind placebo-controlled randomized clinical trial.

Methods

Eighty-four men with low risk prostate cancer were stratified based on self-reported dietary consumption of fish and tomatoes and then randomly assigned to a 3-month intervention of lycopene (n = 29) or fish oil (n = 27) supplementation or placebo (n = 28). Gene expression in morphologically normal prostate tissue was studied at baseline and at 3 months via cDNA microarray analysis. Differential gene expression and pathway analyses were performed to identify genes and pathways modulated by these micronutrients.

Results

Global gene expression analysis revealed no significant individual genes that were associated with high intake of fish or tomato at baseline or after 3 months of supplementation with lycopene or fish oil. However, exploratory pathway analyses of rank-ordered genes (based on p-values not corrected for multiple comparisons) revealed the modulation of androgen and estrogen metabolism in men who routinely consumed more fish (p = 0.029) and tomato (p = 0.008) compared to men who ate less. In addition, modulation of arachidonic acid metabolism (p = 0.01) was observed after 3 months of fish oil supplementation compared with the placebo group; and modulation of nuclear factor (erythroid derived-2) factor 2 or Nrf2-mediated oxidative stress response for either supplement versus placebo (fish oil: p = 0.01, lycopene: p = 0.001).

Conclusions

We did not detect significant individual genes associated with dietary intake and supplementation of lycopene and fish oil. However, exploratory analyses revealed candidate in vivo pathways that may be modulated by these micronutrients.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00402285","term_id":"NCT00402285"}}NCT00402285     

Differences in gene expression between first and third trimester human placenta: a microarray study

Background

The human placenta is a rapidly developing organ that undergoes structural and functional changes throughout the pregnancy. Our objectives were to investigate the differences in global gene expression profile, the expression of imprinted genes and the effect of smoking in first and third trimester normal human placentas.

Materials and Methods

Placental samples were collected from 21 women with uncomplicated pregnancies delivered at term and 16 healthy women undergoing termination of pregnancy at 9–12 weeks gestation. Placental gene expression profile was evaluated by Human Genome Survey Microarray v.2.0 (Applied Biosystems) and real-time polymerase chain reaction.

Results

Almost 25% of the genes spotted on the array (n = 7519) were differentially expressed between first and third trimester placentas. Genes regulating biological processes involved in cell proliferation, cell differentiation and angiogenesis were up-regulated in the first trimester; whereas cell surface receptor mediated signal transduction, G-protein mediated signalling, ion transport, neuronal activities and chemosensory perception were up-regulated in the third trimester. Pathway analysis showed that brain and placenta might share common developmental routes. Principal component analysis based on the expression of 17 imprinted genes showed a clear separation of first and third trimester placentas, indicating that epigenetic modifications occur throughout pregnancy. In smokers, a set of genes encoding oxidoreductases were differentially expressed in both trimesters.

Conclusions

Differences in global gene expression profile between first and third trimester human placenta reflect temporal changes in placental structure and function. Epigenetic rearrangements in the human placenta seem to occur across gestation, indicating the importance of environmental influence in the developing feto-placental unit.     

Expression of human beta-defensins in children with chronic inflammatory bowel disease

Background

Human beta-defensins (hBDs) are antimicrobial peptides known to play a major role in intestinal innate host defence. Altered mucosal expression of hBDs has been suggested to be implicated in chronic inflammatory bowel disease pathogenesis. However, little is known about expression of these peptides in children.

Methods

Intestinal biopsies were obtained from the duodenum (n = 88), terminal ileum (n = 90) and ascending colon (n = 105) of children with Crohn''s disease (n = 26), ulcerative colitis (n = 11) and healthy controls (n = 16). Quantitative real-time (RT) PCR was performed and absolute mRNA copy numbers analyzed for hBD1-3 as well as inflammatory cytokines IL-8 and TNF-alpha.

Results

Significant induction of hBD2 and hBD3 was observed in the inflamed terminal ileum and ascending colon of IBD children. In the ascending colon induction of hBD2 was found to be significantly lower in children with Crohn''s disease compared to ulcerative colitis. A strong correlation was found between inducible defensins hBD2 and 3 and the inflammatory cytokines IL-8 and TNF-alpha, both in the terminal ileum and ascending colon.

Conclusion

Our study demonstrates distinct changes in hBD expression throughout the intestinal tract of children with IBD, lending further support for their potential role in disease pathogenesis.     

MicroRNA signatures in tumor tissue related to angiogenesis in non-small cell lung cancer

Background

Angiogenesis is regarded as a hallmark in cancer development, and anti-angiogenic treatment is presently used in non-small cell lung cancer (NSCLC) patients. MicroRNAs (miRs) are small non-coding, endogenous, single stranded RNAs that regulate gene expression. In this study we aimed to identify significantly altered miRs related to angiogenesis in NSCLC.

Methods

From a large cohort of 335 NSCLC patients, paraffin-embedded samples from 10 patients with a short disease specific survival (DSS), 10 with a long DSS and 10 normal controls were analyzed. The miRs were quantified by microarray hybridization and selected miRs were validated by real-time qPCR. The impacts of different pathways, including angiogenesis, were evaluated by Gene Set Enrichment Analysis (GSEA) derived from Protein ANalysis THrough Evolutionary Relationship (PANTHER). One of the most interesting candidate markers, miR-155, was validated by in situ hybridization (ISH) in the total cohort (n = 335) and correlation analyses with several well-known angiogenic markers were done.

Results

128 miRs were significantly up- or down-regulated; normal versus long DSS (n = 68) and/or normal versus short DSS (n = 63) and/or long versus short DSS (n = 37). The pathway analysis indicates angiogenesis-related miRs to be involved in NSCLC. There were strong significant correlations between the array hybridization and qPCR validation data. The significantly altered angiogenesis-related miRs of high interest were miR-21, miR-106a, miR-126, miR-155, miR-182, miR-210 and miR-424. miR-155 correlated significantly with fibroblast growth factor 2 (FGF2) in the total cohort (r = 0.17, P = 0.002), though most prominent in the subgroup with nodal metastasis (r = 0.34, P<0.001).

Conclusions

Several angiogenesis-related miRs are significantly altered in NSCLC. Further studies to understand their biological functions and explore their clinical relevance are warranted.     

Gene expression profiles in stage I uterine serous carcinoma in comparison to grade 3 and grade 1 stage I endometrioid adenocarcinoma

Background

Endometrial cancer is the most common gynecologic malignancy in the developed countries. Clinical studies have shown that early stage uterine serous carcinoma (USC) has outcomes similar to early stage high grade endometrioid adenocarcinoma (EAC-G3) than to early stage low grade endometrioid adenocarcinoma (EAC-G1). However, little is known about the origin of these different clinical outcomes. This study applied the whole genome expression profiling to explore the expression difference of stage I USC (n = 11) relative to stage I EAC-G3 (n = 11) and stage I EAC-G1 (n = 11), respectively.

Methodology/Principal Finding

We found that the expression difference between USC and EAC-G3, as measured by the number of differentially expressed genes (DEGs), is consistently less than that found between USC and EAC-G1. Pathway enrichment analyses suggested that DEGs specific to USC vs. EAC-G3 are enriched for genes involved in signaling transduction, while DEGs specific to USC vs. EAC-G1 are enriched for genes involved in cell cycle. Gene expression differences for selected DEGs are confirmed by quantitative RT-PCR with a high validation rate.

Conclusion

This data, although preliminary, indicates that stage I USC is genetically similar to stage I EAC-G3 compared to stage I EAC-G1. DEGs identified from this study might provide an insight in to the potential mechanisms that influence the clinical outcome differences between endometrial cancer subtypes. They might also have potential prognostic and therapeutic impacts on patients diagnosed with uterine cancer.     

Urinary podocyte-associated mRNA profile in various stages of diabetic nephropathy

Background

Podocyte injury and subsequent excretion in urine play a crucial role in the pathogenesis and progression of diabetic nephropathy (DN). Quantification of messenger RNA (mRNA) expression in urinary sediment by real-time PCR is emerging as a noninvasive method of screening DN-associated biomarkers. We hypothesized that the urinary mRNA profile of podocyte-associated molecules may provide important clinical insight into the different stages of diabetic nephropathy.

Methods

DN patients (N = 51) and healthy controls (N = 13) were enrolled in this study. DN patients were divided into a normoalbuminuria group (UAE<30 mg/g, n = 17), a microalbuminuria group (UAE 30∼300 mg/g, n = 15), and a macroalbuminuria group (UAE>300 mg/g, n = 19), according to their urinary albumin excretion (UAE). Relative mRNA abundance of synaptopodin, podocalyxin, CD2-AP, α-actin4, and podocin were quantified, and correlations between target mRNAs and clinical parameters were examined.

Results

The urinary mRNA levels of all genes studied were significantly higher in the DN group compared with controls (p<0.05), and mRNA levels increased with DN progression. Urinary mRNA levels of all target genes positively correlated with both UAE and BUN. The expression of podocalyxin, CD2-AP, α-actin4, and podocin mRNA correlated with serum creatinine (r = 0.457, p = 0.001; r = 0.329, p = 0.01; r = 0.286, p = 0.021; r = 0.357, p = 0.006, respectively). Furthermore, podocalyxin mRNA was found to negatively correlate with eGFR (r = −0.349, p = 0.01).

Conclusion

The urinary mRNA profiles of synaptopodin, podocalyxin, CD2-AP, α-actin4, and podocin were found to increase with the progression of DN, which suggested that quantification of podocyte-associated molecules will be useful biomarkers of DN.     

Developing a gene expression model for predicting ventilator-associated pneumonia in trauma patients: a pilot study

Background

Ventilator-associated pneumonia (VAP) carries significant mortality and morbidity. Predicting which patients will become infected could lead to measures to reduce the incidence of VAP.

Methodology/Principal Findings

The goal was to begin constructing a model for VAP prediction in critically-injured trauma patients, and to identify differentially expressed genes in patients who go on to develop VAP compared to similar patients who do not. Gene expression profiles of lipopolysaccharide stimulated blood cells in critically injured trauma patients that went on to develop ventilator-associated pneumonia (n = 10) was compared to those that never developed the infection (n = 10). Eight hundred and ten genes were differentially expressed between the two groups (ANOVA, P<0.05) and further analyzed by hierarchical clustering and principal component analysis. Functional analysis using Gene Ontology and KEGG classifications revealed enrichment in multiple categories including regulation of protein translation, regulation of protease activity, and response to bacterial infection. A logistic regression model was developed that accurately predicted critically-injured trauma patients that went on to develop VAP (VAP+) and those that did not (VAP−). Five genes (PIK3R3, ATP2A1, PI3, ADAM8, and HCN4) were common to all top 20 significant genes that were identified from all independent training sets in the cross validation. Hierarchical clustering using these five genes accurately categorized 95% of patients and PCA visualization demonstrated two discernable groups (VAP+ and VAP−).

Conclusions/Significance

A logistic regression model using cross-validation accurately predicted patients that developed ventilator-associated pneumonia and should now be tested on a larger cohort of trauma patients.     

Soluble serum CD81 is elevated in patients with chronic hepatitis C and correlates with alanine aminotransferase serum activity

Aim

Cellular CD81 is a well characterized hepatitis C virus (HCV) entry factor, while the relevance of soluble exosomal CD81 in HCV pathogenesis is poorly defined. We performed a case-control study to investigate whether soluble CD81 in the exosomal serum fraction is associated with HCV replication and inflammatory activity.

Patients and Methods

Four cohorts were investigated, patients with chronic hepatitis C (n = 37), patients with chronic HCV infection and persistently normal ALT levels (n = 24), patients with long term sustained virologic response (SVR, n = 7), and healthy volunteers (n = 23). Concentration of soluble CD81 was assessed semi-quantitatively after differential centrifugation ranging from 200 g to 100,000 g in the fifth centrifugation fraction by immunoblotting and densitometry.

Results

Soluble CD81 was increased in patients with chronic hepatitis C compared to healthy subjects (p = 0.03) and cured patients (p = 0.017). Patients with chronic HCV infection and persistently normal ALT levels and patients with long term SVR had similar soluble CD81 levels as healthy controls (p>0.2). Overall, soluble CD81 levels were associated with ALT levels (r = 0.334, p = 0.016) and severe liver fibrosis (p = 0.027).

Conclusion

CD81 is increased in the exosomal serum fraction in patients with chronic hepatitis C and appears to be associated with inflammatory activity and severity of fibrosis.     

Clinical, biological and genetic analysis of anorchia in 26 boys

Background

Anorchia is defined as the absence of testes in a 46,XY individual with a male phenotype. The cause is unknown.

Methods

We evaluated the clinical and biological presentation, and family histories of 26 boys with anorchia, and sequenced their SRY, NR5A1, INSL3, MAMLD1 genes and the T222P variant for LGR8.

Results

No patient had any associated congenital anomaly. At birth, testes were palpable bilaterally or unilaterally in 13 cases and not in 7; one patient presented with bilateral testicular torsion immediately after birth. The basal plasma concentrations of anti-Müllerian hormone (AMH, n = 15), inhibin B (n = 7) and testosterone (n = 19) were very low or undetectable in all the patients evaluated, as were the increases in testosterone after human chorionic gonadotropin (hCG, n = 12). The basal plasma concentrations of follicle stimulating hormone (FSH) were increased in 20/25, as was that of luteinising hormone in 10/22 cases. Family members of 7/26 cases had histories of primary ovarian failure in the mother (n = 2), or sister 46,XX, together with fetal malformations of the only boy with microphallus and secondary foot edema (n = 1), secondary infertility in the father (n = 2), or cryptorchidism in first cousins (n = 2). The sequences of all the genes studied were normal.

Conclusion

Undetectable plasma concentrations of AMH and inhibin B and an elevated plasma FSH, together with 46,XY complement are sufficient for diagnosis of anorchia. The hCG test is unnecessary. NR5A1 and other genes implicated in gonadal development and testicle descent were not mutated, which suggests that other genes involved in these developments contribute to the phenotypes.     

Beta defensin-2 is reduced in central but not in distal airways of smoker COPD patients

Background

Altered pulmonary defenses in chronic obstructive pulmonary disease (COPD) may promote distal airways bacterial colonization. The expression/activation of Toll Like receptors (TLR) and beta 2 defensin (HBD2) release by epithelial cells crucially affect pulmonary defence mechanisms.

Methods

The epithelial expression of TLR4 and of HBD2 was assessed in surgical specimens from current smokers COPD (s-COPD; n = 17), ex-smokers COPD (ex-s-COPD; n = 8), smokers without COPD (S; n = 12), and from non-smoker non-COPD subjects (C; n = 13).

Results

In distal airways, s-COPD highly expressed TLR4 and HBD2. In central airways, S and s-COPD showed increased TLR4 expression. Lower HBD2 expression was observed in central airways of s-COPD when compared to S and to ex-s-COPD. s-COPD had a reduced HBD2 gene expression as demonstrated by real-time PCR on micro-dissected bronchial epithelial cells. Furthermore, HBD2 expression positively correlated with FEV1/FVC ratio and inversely correlated with the cigarette smoke exposure. In a bronchial epithelial cell line (16 HBE) IL-1β significantly induced the HBD2 mRNA expression and cigarette smoke extracts significantly counteracted this IL-1 mediated effect reducing both the activation of NFkB pathway and the interaction between NFkB and HBD2 promoter.

Conclusions

This study provides new insights on the possible mechanisms involved in the alteration of innate immunity mechanisms in COPD.     

Fatty acid binding protein 1 is related with development of aspirin-exacerbated respiratory disease

Background

Aspirin-exacerbated respiratory disease (AERD) refers to the development of bronchoconstriction in asthmatics following the ingestion of aspirin. Although alterations in eicosanoid metabolites play a role in AERD, other immune or inflammatory mechanisms may be involved. We aimed to identify proteins that were differentially expressed in nasal polyps between patients with AERD and aspirin-tolerant asthma (ATA).

Methodology/Principal Findings

Two-dimensional electrophoresis was adopted for differential display proteomics. Proteins were identified by liquid chromatography-tandem mass spectrometry (LC-MS). Western blotting and immunohistochemical staining were performed to compare the amount of fatty acid-binding protein 1 (FABP1) in the nasal polyps of patients with AERD and ATA. Fifteen proteins were significantly up- (seven spots) or down-regulated in the nasal polyps of patients with AERD (n = 5) compared to those with ATA (n = 8). LC-MS revealed an increase in seven proteins expression and a decrease in eight proteins expression in patients with AERD compared to those with ATA (P = 0.003–0.045). FABP1-expression based on immunoblotting and immunohistochemical analysis was significantly higher in the nasal polyps of patients with AERD compared to that in patients with ATA. FABP1 was observed in epithelial, eosinophils, macrophages, and the smooth-muscle cells of blood vessels in the polyps.

Conclusions/Significance

Our results indicate that alterations in 15 proteins, including FABP1, may be related to the development of AERD.     

Comparative profiling of primary colorectal carcinomas and liver metastases identifies LEF1 as a prognostic biomarker

Purpose

We sought to identify genes of clinical significance to predict survival and the risk for colorectal liver metastasis (CLM), the most common site of metastasis from colorectal cancer (CRC).

Patients and Methods

We profiled gene expression in 31 specimens from primary CRC and 32 unmatched specimens of CLM, and performed Significance Analysis of Microarrays (SAM) to identify genes differentially expressed between these two groups. To characterize the clinical relevance of two highly-ranked differentially-expressed genes, we analyzed the expression of secreted phosphoprotein 1 (SPP1 or osteopontin) and lymphoid enhancer factor-1 (LEF1) by immunohistochemistry using a tissue microarray (TMA) representing an independent set of 154 patients with primary CRC.

Results

Supervised analysis using SAM identified 963 genes with significantly higher expression in CLM compared to primary CRC, with a false discovery rate of <0.5%. TMA analysis showed SPP1 and LEF1 protein overexpression in 60% and 44% of CRC cases, respectively. Subsequent occurrence of CLM was significantly correlated with the overexpression of LEF1 (chi-square p = 0.042), but not SPP1 (p = 0.14). Kaplan Meier analysis revealed significantly worse survival in patients with overexpression of LEF1 (p<0.01), but not SPP1 (p = 0.11). Both univariate and multivariate analyses identified stage (p<0.0001) and LEF1 overexpression (p<0.05) as important prognostic markers, but not tumor grade or SPP1.

Conclusion

Among genes differentially expressed between CLM and primary CRC, we demonstrate overexpression of LEF1 in primary CRC to be a prognostic factor for poor survival and increased risk for liver metastasis.