Genome-wide DNA Microarray Analysis Research Articles

Transcriptional changes involved in inhibition of biofilm formation by ε-polylysine in Salmonella Typhimurium.

The pathogenicity of Salmonella Typhimurium, a foodborne pathogen, is mainly attributed to its ability to form biofilm on food contact surfaces. ε-polylysine, a polymer of positively charged lysine, is reported to inhibit biofilm formation of both gram-positive and gram-negative bacteria. To elucidate the mechanism underlying ε-polylysine-mediated inhibition of biofilm formation, the transcriptional profiles of ε-polylysine-treated and untreated Salmonella Typhimurium cells were comparatively analysed. The genome-wide DNA microarray analysis was performed using Salmonella Typhimurium incubated with 0.001% ε-polylysine in 0.1% Bacto Soytone at 30°C for 2h. The expression levels of genes involved in curli amyloid fibres and cellulose production, quorum sensing, and flagellar motility were downregulated, whereas those of genes associated with colanic acid synthesis were upregulated after treatment with ε-polylysine. The microarray results were validated by quantitative real-time polymerase chain reaction (qRT-PCR). Furthermore, treatment with ε-polylysine decreased the production of colanic acid in Salmonella Typhimurium. The findings of this study improved our understanding of the mechanisms underlying ε-polylysine-mediated biofilm inhibition and may contribute to the development of new disinfectants to control biofilm during food manufacturing and storage.

Comparison of gene expression profiles in drug-withdrawn rats

The aim of the present study was to compare gene expression profiles in animals treated with abusable drugs. Wistar rats were treated with saline, morphine, chloral hydrate, or pentobarbital twice daily for 14 days. To induce neurotoxicity, a group of rats was treated with kainic acid. 24 hr after withdrawal, and a genome-wide DNA microarray analysis was conducted with the striata. A total of 57 gene expression patterns were altered in the three test groups; 35 of the 157 genes were selected when compared with the kainic acid treated group. Twenty-two of the 24 genes were functionally described in the gene library, and many were involved in long-term potentiation (LTP) and cell adhesion. The results were confirmed by quantitative reverse transcription-polymerase chain reaction analysis. Taken together, the selected genes may subserve identification of physical dependence biomarkers and contribute to determining the molecular mechanisms underlying physical dependence.

DNA Microarray Highlights Nrf2-Mediated Neuron Protection Targeted by Wasabi-Derived Isothiocyanates in IMR-32 Cells

6-(Methylsulfinyl)hexyl isothiocyanate (6-MSITC), 6-(methylthio)hexyl isothiocyanate (6-MTITC), and 4-(methylsulfinyl)butyl isothiocyanate (4-MSITC) are isothiocyanate (ITC) bioactive compounds from Japanese Wasabi. Previous in vivo studies highlighted the neuroprotective potential of ITCs since ITCs enhance the production of antioxidant-related enzymes. Thus, in this present study, a genome-wide DNA microarray analysis was designed to profile gene expression changes in a neuron cell line, IMR-32, stimulated by these ITCs. Among these ITCs, 6-MSITC caused the expression changes of most genes (263), of which 100 genes were upregulated and 163 genes were downregulated. Gene categorization showed that most of the differentially expressed genes are involved in oxidative stress response, and pathway analysis further revealed that Nrf2-mediated oxidative stress pathway is the top of the ITC-modulated signaling pathway. Finally, real-time polymerase chain reaction (PCR) and Western blotting confirmed the gene expression and protein products of the major targets by ITCs. Taken together, Wasabi-derived ITCs might target the Nrf2-mediated oxidative stress pathway to exert neuroprotective effects.

The Anxiolytic and Antidepressant-like Effects of Testosterone and Estrogen in Gonadectomized Male Rats

BackgroundWhile the influence of testosterone levels on vulnerability to affective disorders is not straightforward, research suggests this hormone may confer some degree of resiliency in men. We recently demonstrated a role for the dentate gyrus in mediating testosterone’s protective effects on depressive-like behavior in gonadectomized male rats. Here, testosterone may exert its effects through androgen receptor-mediated mechanisms or via local aromatization to estradiol. MethodsGonadectomized male rats were implanted with a placebo, testosterone, or estradiol pellet, and subsequent protective anxiolytic- and antidepressant-like effects of testosterone and its aromatized metabolite, estradiol, were then investigated in the open field and sucrose preference tests, respectively. Moreover, their influence on gene expression in the hippocampus was analyzed by genome-wide complementary DNA microarray analysis. Finally, the contribution of testosterone’s aromatization within the dentate gyrus was assessed by local infusion of the aromatase inhibitor fadrozole, whose efficacy was confirmed by liquid chromatography-tandem mass spectrometry. ResultsBoth hormones had antidepressant-like effects associated with a substantial overlap in transcriptional regulation, particularly in synaptic plasticity- and mitogen-activated protein kinase pathway-related genes. Further, chronic aromatase inhibition within the dentate gyrus blocked the protective effects of testosterone. ConclusionsBoth testosterone and estradiol exhibit anxiolytic- and antidepressant-like effects in gonadectomized male rats, while similarly regulating critical mediators of these behaviors, suggesting common underlying mechanisms. Accordingly, we demonstrated that testosterone’s protective effects are mediated, in part, by its aromatization in the dentate gyrus. These findings thus provide further insight into a role for estradiol in mediating the protective anxiolytic- and antidepressant-like effects of testosterone.

FRI0007 Prediction of Therapeutic Responses to TOCILIZUMAB in Patients with Rheumatoid Arthritis Using Biomarkers Identified by Genome-Wide DNA Microarray Analysis in Peripheral Blood Mononuclear Cells

BackgroundAlthough the overall response rate of rheumatoid arthritis (RA) to tocilizumab is high, improvement of synovitis in patients treated with tocilizumab is frequently slow as compared with that in patients...

Prediction of Therapeutic Responses to Tocilizumab in Patients With Rheumatoid Arthritis: Biomarkers Identified by Analysis of Gene Expression in Peripheral Blood Mononuclear Cells Using Genome‐Wide DNA Microarray

The aim of this prospective multicenter study was to identify biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with rheumatoid arthritis (RA). We recruited patients with RA who were treated with tocilizumab for the first time, and determined therapeutic responses at 6 months. In the training cohort (n = 40), gene expression in peripheral blood mononuclear cells (PBMCs) at baseline was analyzed using genome-wide DNA microarray, with 41,000 probes derived from 19,416 genes. In the validation cohort (n = 20), expression levels of the candidate genes in PBMCs at baseline were determined using real-time quantitative polymerase chain reaction (qPCR) analysis. We identified 68 DNA microarray probes that showed significant differences in signal intensity between nonresponders and responders in the training cohort. Nineteen putative genes were selected, and a significant correlation between the DNA microarray signal intensity and the qPCR relative expression was confirmed in 15 genes. In the validation cohort, a significant difference in relative expression between nonresponders and responders was reproduced for 3 type I interferon response genes (IFI6, MX2, and OASL) and MT1G. Receiver operating characteristic curve analysis of models incorporating these genes showed that the maximum area under the curve was 0.947 in predicting a moderate or good response to tocilizumab in the validation cohort. Using genome-wide DNA microarray analyses, we identified candidate biomarkers that can be used to predict therapeutic responses to tocilizumab in patients with RA. These findings suggest that type I interferon signaling and metallothioneins are involved in the pathophysiology of RA.

Whole Blood Transcriptomics and Urinary Metabolomics to Define Adaptive Biochemical Pathways of High-Intensity Exercise in 50-60 Year Old Masters Athletes

Exercise is beneficial for a variety of age-related disorders. However, the molecular mechanisms mediating the beneficial adaptations to exercise in older adults are not well understood. The aim of the current study was to utilize a dual approach to characterize the genetic and metabolic adaptive pathways altered by exercise in veteran athletes and age-matched untrained individuals. Two groups of 50–60 year old males: competitive cyclists (athletes, n = 9; VO2peak 59.1±5.2 ml·kg−1·min−1; peak aerobic power 383±39 W) and untrained, minimally active individuals (controls, n = 8; VO2peak 35.9±9.7 ml·kg−1·min−1; peak aerobic power 230±57 W) were examined. All participants completed an acute bout of submaximal endurance exercise, and blood and urine samples pre- and post-exercise were analyzed for gene expression and metabolic changes utilizing genome-wide DNA microarray analysis and NMR spectroscopy-based metabolomics, respectively. Our results indicate distinct differences in gene and metabolite expression involving energy metabolism, lipids, insulin signaling and cardiovascular function between the two groups. These findings may lead to new insights into beneficial signaling pathways of healthy aging and help identify surrogate markers for monitoring exercise and training load.

Pharmacological Inhibition of Cochlear Mitochondrial Respiratory Chain Induces Secondary Inflammation in the Lateral Wall: A Potential Therapeutic Target for Sensorineural Hearing Loss

Cochlear lateral wall has recently been reported as a common site of inflammation, yet precise molecular mechanisms of the inflammatory responses remain elucidated. The present study examined the inflammatory responses in the lateral wall following acute mitochondrial dysfunction induced by a mitochondrial toxin, 3-nitropropionic acid (3-NP). Reverse-transcription (RT)-PCR revealed increases in the expression of the proinflammatory cytokines interleukin (IL)-1β and IL-6. Immunohistochemistry showed an increase in the number of activated cochlear macrophages in the lateral wall, which were in close proximity to IL-6-expressing cells. A genome-wide DNA microarray analysis of the lateral wall revealed that 35% and 60% of the genes showing >2-fold upregulation at 1 d and 3 d post-3-NP administration, respectively, were inflammatory genes, including CC- and CXC-type chemokine genes. High expression of CCL-1, 2, and 3 at 1 d, and of CCL-1, 2, 3, 4, and 5, CCR-2 and 5, and CX3CR1 at 3 d post-3-NP administration, coupled with no change in the level of CX3CL1 expression suggested that macrophages and monocytes may be involved in the inflammatory response to 3-NP-mediated injury. Quantitative (q)RT-PCR showed a transient induction of IL-1β and IL-6 expression within 24 h of 3-NP-mediated injury, followed by sustained expression of the chemoattractants, CCL-2, 4 and 5, up until 7 d after injury. The expression of CCL-2 and IL-6 was higher in animals showing permanent hearing impairment than in those showing temporary hearing impairment, suggesting that these inflammatory responses may be detrimental to hearing recovery. The present findings suggest that acute mitochondrial dysfunction induces secondary inflammatory responses in the lateral wall of the cochlear and that the IL-6/CCL-2 inflammatory pathway is involved in monocyte activation. Therefore, these secondary inflammatory responses may be a potential post-insult therapeutic target for treatments aimed at preventing the damage caused by acute mitochondrial dysfunction in the cochlear lateral wall.

Silencing diacylglycerol kinase-theta expression reduces steroid hormone biosynthesis and cholesterol metabolism in human adrenocortical cells

Diacylglycerol kinase theta (DGKθ) plays a pivotal role in regulating adrenocortical steroidogenesis by synthesizing the ligand for the nuclear receptor steroidogenic factor 1 (SF1). In response to activation of the cAMP signaling cascade nuclear DGK activity is rapidly increased, facilitating PA-mediated, SF1-dependent transcription of genes required for cortisol and dehydroepiandrosterone (DHEA) biosynthesis. Based on our previous work identifying DGKθ as the enzyme that produces the agonist for SF1, we generated a tetracycline-inducible H295R stable cell line to express a short hairpin RNA (shRNA) against DGKθ and characterized the effect of silencing DGKθ on adrenocortical gene expression. Genome-wide DNA microarray analysis revealed that silencing DGKθ expression alters the expression of multiple genes, including steroidogenic genes, nuclear receptors and genes involved in sphingolipid, phospholipid and cholesterol metabolism. Interestingly, the expression of sterol regulatory element binding proteins (SREBPs) was also suppressed. Consistent with the suppression of SREBPs, we observed a down-regulation of multiple SREBP target genes, including 3-hydroxy-3-methylglutary coenzyme A reductase (HMG-CoA red) and CYP51, concomitant with a decrease in cellular cholesterol. DGKθ knockdown cells exhibited a reduced capacity to metabolize PA, with a down-regulation of lipin and phospholipase D (PLD) isoforms. In contrast, suppression of DGKθ increased the expression of several genes in the sphingolipid metabolic pathway, including acid ceramidase (ASAH1) and sphingosine kinases (SPHK). In summary, these data demonstrate that DGKθ plays an important role in steroid hormone production in human adrenocortical cells.

Enhancing E. coli isobutanol tolerance through engineering its global transcription factor cAMP receptor protein (CRP)

The limited isobutanol tolerance of Escherichia coli is a major drawback during fermentative isobutanol production. Different from classical strain engineering approaches, this work was initiated to improve E. coli isobutanol tolerance from its transcriptional level by engineering its global transcription factor cAMP receptor protein (CRP). Random mutagenesis libraries were generated by error-prone PCR of crp, and the libraries were subjected to isobutanol stress for selection. Variant IB2 (S179P, H199R) was isolated and exhibited much better growth (0.18 h(-1) ) than the control (0.05 h(-1) ) in 1.2% (v/v) isobutanol (9.6 g/L). Genome-wide DNA microarray analysis revealed that 58 and 308 genes in IB2 had differential expression (>2-fold, p < 0.05) in the absence and presence of 1% (v/v) isobutanol, respectively. When challenged with isobutanol, genes related to acid resistance (gadABCE, hdeABD), nitrate reduction (narUZYWV), flagella and fimbrial activity (lfhA, yehB, ycgR, fimCDF), and sulfate reduction and transportation (cysIJH, cysC, cysN) were the major functional groups that were up-regulated, whereas most of the down-regulated genes were enzyme (tnaA) and transporters (proVWX, manXYZ). As demonstrated by single-gene knockout experiments, gadX, nirB, rhaS, hdeB, and ybaS were found associated with strain isobutanol resistance. The intracellular reactive oxygen species (ROS) level in IB2 was only half of that of the control when facing stress, indicating that IB2 can withstand toxic isobutanol much better than the control.

Enhancing E. coli Tolerance towards Oxidative Stress via Engineering Its Global Regulator cAMP Receptor Protein (CRP)

Oxidative damage to microbial hosts often occurs under stressful conditions during bioprocessing. Classical strain engineering approaches are usually both time-consuming and labor intensive. Here, we aim to improve E. coli performance under oxidative stress via engineering its global regulator cAMP receptor protein (CRP), which can directly or indirectly regulate redox-sensing regulators SoxR and OxyR, and other ∼400 genes in E. coli. Error-prone PCR technique was employed to introduce modifications to CRP, and three mutants (OM1∼OM3) were identified with improved tolerance via H2O2 enrichment selection. The best mutant OM3 could grow in 12 mM H2O2 with the growth rate of 0.6 h−1, whereas the growth of wild type was completely inhibited at this H2O2 concentration. OM3 also elicited enhanced thermotolerance at 48°C as well as resistance against cumene hydroperoxide. The investigation about intracellular reactive oxygen species (ROS), which determines cell viability, indicated that the accumulation of ROS in OM3 was always lower than in WT with or without H2O2 treatment. Genome-wide DNA microarray analysis has shown not only CRP-regulated genes have demonstrated great transcriptional level changes (up to 8.9-fold), but also RpoS- and OxyR-regulated genes (up to 7.7-fold). qRT-PCR data and enzyme activity assay suggested that catalase (katE) could be a major antioxidant enzyme in OM3 instead of alkyl hydroperoxide reductase or superoxide dismutase. To our knowledge, this is the first work on improving E. coli oxidative stress resistance by reframing its transcription machinery through its native global regulator. The positive outcome of this approach may suggest that engineering CRP can be successfully implemented as an efficient strain engineering alternative for E. coli.

Identification and Functional Analysis of Epigenetically Silenced MicroRNAs in Colorectal Cancer Cells

Abnormal microRNA (miRNA) expression has been linked to the development and progression of several human cancers, and such dysregulation can result from aberrant DNA methylation. While a small number of miRNAs is known to be regulated by DNA methylation, we postulated that such epigenetic regulation is more prevalent. By combining MBD-isolated Genome Sequencing (MiGS) to evaluate genome-wide DNA methylation patterns and microarray analysis to determine miRNA expression levels, we systematically searched for candidate miRNAs regulated by DNA methylation in colorectal cancer cell lines. We found 64 miRNAs to be robustly methylated in HCT116 cells; eighteen of them were located in imprinting regions or already reported to be regulated by DNA methylation. For the remaining 46 miRNAs, expression levels of 18 were consistent with their DNA methylation status. Finally, 8 miRNAs were up-regulated by 5-aza-2′-deoxycytidine treatment and identified to be novel miRNAs regulated by DNA methylation. Moreover, we demonstrated the functional relevance of these epigenetically silenced miRNAs by ectopically expressing select candidates, which resulted in inhibition of growth and migration of cancer cells. In addition to reporting these findings, our study also provides a reliable, systematic strategy to identify DNA methylation-regulated miRNAs by combining DNA methylation profiles and expression data.

Altered gene regulation and synaptic morphology in Drosophila learning and memory mutants

Genetic studies in Drosophila have revealed two separable long-term memory pathways defined as anesthesia-resistant memory (ARM) and long-lasting long-term memory (LLTM). ARM is disrupted in radish (rsh) mutants, whereas LLTM requires CREB-dependent protein synthesis. Although the downstream effectors of ARM and LLTM are distinct, pathways leading to these forms of memory may share the cAMP cascade critical for associative learning. Dunce, which encodes a cAMP-specific phosphodiesterase, and rutabaga, which encodes an adenylyl cyclase, both disrupt short-term memory. Amnesiac encodes a pituitary adenylyl cyclase-activating peptide homolog and is required for middle-term memory. Here, we demonstrate that the Radish protein localizes to the cytoplasm and nucleus and is a PKA phosphorylation target in vitro. To characterize how these plasticity pathways may manifest at the synaptic level, we assayed synaptic connectivity and performed an expression analysis to detect altered transcriptional networks in rutabaga, dunce, amnesiac, and radish mutants. All four mutants disrupt specific aspects of synaptic connectivity at larval neuromuscular junctions (NMJs). Genome-wide DNA microarray analysis revealed ∼375 transcripts that are altered in these mutants, suggesting defects in multiple neuronal signaling pathways. In particular, the transcriptional target Lapsyn, which encodes a leucine-rich repeat cell adhesion protein, localizes to synapses and regulates synaptic growth. This analysis provides insights into the Radish-dependent ARM pathway and novel transcriptional targets that may contribute to memory processing in Drosophila.

A proteomic and transcriptional view of acidogenic and solventogenic steady-state cells of Clostridium acetobutylicum in a chemostat culture

The complex changes in the life cycle of Clostridium acetobutylicum, a promising biofuel producer, are not well understood. During exponential growth, sugars are fermented to acetate and butyrate, and in the transition phase, the metabolism switches to the production of the solvents acetone and butanol accompanied by the initiation of endospore formation. Using phosphate-limited chemostat cultures at pH 5.7, C. acetobutylicum was kept at a steady state of acidogenic metabolism, whereas at pH 4.5, the cells showed stable solvent production without sporulation. Novel proteome reference maps of cytosolic proteins from both acidogenesis and solventogenesis with a high degree of reproducibility were generated. Yielding a 21% coverage, 15 protein spots were specifically assigned to the acidogenic phase, and 29 protein spots exhibited a significantly higher abundance in the solventogenic phase. Besides well-known metabolic proteins, unexpected proteins were also identified. Among these, the two proteins CAP0036 and CAP0037 of unknown function were found as major striking indicator proteins in acidogenic cells. Proteome data were confirmed by genome-wide DNA microarray analyses of the identical cultures. Thus, a first systematic study of acidogenic and solventogenic chemostat cultures is presented, and similarities as well as differences to previous studies of batch cultures are discussed.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-010-2741-x) contains supplementary material, which is available to authorized users.

Stress sensors and signal transducers in cyanobacteria.

In living cells, the perception of environmental stress and the subsequent transduction of stress signals are primary events in the acclimation to changes in the environment. Some molecular sensors and transducers of environmental stress cannot be identified by traditional and conventional methods. Based on genomic information, a systematic approach has been applied to the solution of this problem in cyanobacteria, involving mutagenesis of potential sensors and signal transducers in combination with DNA microarray analyses for the genome-wide expression of genes. Forty-five genes for the histidine kinases (Hiks), 12 genes for serine-threonine protein kinases (Spks), 42 genes for response regulators (Rres), seven genes for RNA polymerase sigma factors, and nearly 70 genes for transcription factors have been successfully inactivated by targeted mutagenesis in the unicellular cyanobacterium Synechocystis sp. PCC 6803. Screening of mutant libraries by genome-wide DNA microarray analysis under various stress and non-stress conditions has allowed identification of proteins that perceive and transduce signals of environmental stress. Here we summarize recent progress in the identification of sensory and regulatory systems, including Hiks, Rres, Spks, sigma factors, transcription factors, and the role of genomic DNA supercoiling in the regulation of the responses of cyanobacterial cells to various types of stress.

Gene expression profiling following short-term and long-term morphine exposure in mice uncovers genes involved in food intake

Addictive drugs including opioids activate signal transduction pathways that regulate gene expression in the brain. However, changes in CNS gene expression following morphine exposure are poorly understood. We determined changes in gene expression following short- and long-term morphine treatment in the hypothalamus and pituitary using genome-wide DNA microarray analysis and confirmed those alterations in gene expression by real-time reverse transcriptase polymerase chain reaction (RT-PCR) analysis. In the hypothalamus, short-term morphine administration up-regulated (at least twofold) 39 genes and down-regulated six genes. Long-term morphine treatment up-regulated 35 genes and down-regulated 51 genes. In the pituitary, short-term morphine administration up-regulated 110 genes and down-regulated 29 genes. Long-term morphine treatment up-regulated 85 genes and down-regulated 37 pituitary genes. Microarray analysis uncovered several genes involved in food intake (neuropeptide Y, agouti-related protein, and cocaine and amphetamine-regulated transcript) whose expression was strongly altered by morphine exposure in either the hypothalamus or pituitary. Subsequent RT-PCR analysis confirmed similar regulation in expression of these genes in the hypothalamus and pituitary. Finally, we found functional correlation between morphine-induced alterations in food intake and regulation of genes involved in this process. Changes in genes related to food intake may uncover new pathways related to some of the physiological effects of opioids.

Eviprostat suppresses proinflammatory gene expression in the prostate of rats with partial bladder-outlet obstruction: A genome-wide DNA microarray analysis

Prostatic inflammation plays a role in the progression of benign prostatic hyperplasia (BPH). Eviprostat is an antioxidant, antiinflammatory phytotherapeutic agent widely used to treat lower urinary tract symptoms in BPH. Because Eviprostat is a mixture of compounds from multiple natural sources, however, its mechanism of action has been difficult to investigate. Here, we describe the use of oligonucleotide microarrays to investigate changes in gene expression in the prostate of rats with surgically induced partial bladder-outlet obstruction and the effect of Eviprostat on those changes. Several dozen proinflammatory genes were activated in obstructed rats, including cytokine, arachidonic acid cascade enzyme, Toll-like receptor (TLR), and transcription factor genes, and their expression was suppressed by Eviprostat. Pathway analysis revealed that several proinflammatory pathways were activated, including cytokine and TLR signaling pathways. The differential expression of selected genes was verified by real-time reverse-transcriptase polymerase chain reaction. Our findings suggest that prostate inflammation in our rat model of partial bladder-outlet obstruction is related to the increased expression of nuclear factor κB (NF-κB) and the induction of proinflammatory cytokines, and that Eviprostat suppresses their expression at the transcriptional level. The prostate inflammation seen in BPH and the clinical benefits of Eviprostat may be similarly explained.

Aberrant gene expression profile in a mouse model of endometriosis mirrors that observed in women

To define the altered gene expression profile of endometriotic lesions in a mouse model of surgically induced endometriosis. Autologous experimental mouse model. Medical school department. Adult C57Bl6 mice. Endometriosis was surgically induced by autotransplantation of uterine tissue to the intestinal mesentery. Endometriotic lesions and eutopic uteri were recovered at 3 or 29 days after induction. Altered gene expression was measured in the endometriotic lesion relative to the eutopic uterus by genome-wide complementary DNA microarray analysis and was confirmed by real-time reverse transcriptase-polymerase chain reaction for six genes. Relevant categories of altered genes were identified using gene ontology analysis to determine groups of genes enriched for altered expression. The expression of 479 and 114 genes was altered in the endometriotic lesion compared with the eutopic uterus at 3 or 29 days after induction, respectively. Gene ontology enrichment analysis revealed that genes associated with the extracellular matrix, cell adhesions, immune function, cell growth, and angiogenesis were altered in the endometriotic lesion compared with the eutopic uterus. According to gene expression analysis, the mouse model of surgically induced endometriosis is a good model for studying the pathophysiology and treatment of endometriosis.

PGC-1α Expression Decreases in the Alzheimer Disease Brain as a Function of Dementia

To explore mechanisms through which altered peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC-1alpha) expression may influence Alzheimer disease (AD) amyloid neuropathology and to test the hypothesis that promotion of PGC-1alpha expression in neurons might be developed as a novel therapeutic strategy in AD. Case-control. Patients Human postmortem brain (hippocampal formation) samples from AD cases and age-matched non-AD cases. Using genome-wide complementary DNA microarray analysis, we found that PGC-1alpha messenger RNA expression was significantly decreased as a function of progression of clinical dementia in the AD brain. Following confirmatory real-time polymerase chain reaction assay, we continued to explore the role of PGC-1alpha in clinical dementia and found that PGC-1alpha protein content was negatively associated with both AD-type neuritic plaque pathology and beta-amyloid (Abeta)(X-42) contents. Moreover, we found that the predicted elevation of amyloidogenic Abeta(1-42) and Abeta(1-40) peptide accumulation in embryonic cortico-hippocampal neurons derived from Tg2576 AD mice under hyperglycemic conditions (glucose level, 182-273 mg/dL) coincided with a dose-dependent attenuation in PGC-1alpha expression. Most importantly, we found that the reconstitution of exogenous PGC-1alpha expression in Tg2576 neurons attenuated the hyperglycemic-mediated beta-amyloidogenesis through mechanisms involving the promotion of the "nonamyloidogenic" alpha-secretase processing of amyloid precursor protein through the attenuation of the forkheadlike transcription factor 1 (FoxO3a) expression. Therapeutic preservation of neuronal PGC-1alpha expression promotes the nonamyloidogenic processing of amyloid precursor protein precluding the generation of amyloidogenic Abeta peptides.

Delineation of VEGF-regulated genes and functions in the cervix of pregnant rodents by DNA microarray analysis

BackgroundVEGF-regulated genes in the cervices of pregnant and non-pregnant rodents (rats and mice) were delineated by DNA microarray and Real Time PCR, after locally altering levels of or action of VEGF using VEGF agents, namely siRNA, VEGF receptor antagonist and mouse VEGF recombinant protein.MethodsTissues were analyzed by genome-wide DNA microarray analysis, Real-time and gel-based PCR, and SEM, to decipher VEGF function during cervical remodeling. Data were analyzed by EASE score (microarray) and ANOVA (Real Time PCR) followed by Scheffe's F-test for multiple comparisons.ResultsOf the 30,000 genes analyzed, about 4,200 genes were altered in expression by VEGF, i.e., expression of about 2,400 and 1,700 genes were down- and up-regulated, respectively. Based on EASE score, i.e., grouping of genes according to their biological process, cell component and molecular functions, a number of vascular- and non-vascular-related processes were found to be regulated by VEGF in the cervix, including immune response (including inflammatory), cell proliferation, protein kinase activity, and cell adhesion molecule activity. Of interest, mRNA levels of a select group of genes, known to or with potential to influence cervical remodeling were altered. For example, real time PCR analysis showed that levels of VCAM-1, a key molecule in leukocyte recruitment, endothelial adhesion, and subsequent trans-endothelial migration, were elevated about 10 folds by VEGF. Further, VEGF agents also altered mRNA levels of decorin, which is involved in cervical collagen fibrillogenesis, and expression of eNO, PLC and PKC mRNA, critical downstream mediators of VEGF. Of note, we show that VEGF may regulate cervical epithelial proliferation, as revealed by SEM.ConclusionThese data are important in that they shed new insights in VEGF's possible roles and mechanisms in cervical events near-term, including cervical remodeling.