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1.  Changes in mouse cognition and hippocampal gene expression observed in a mild physical- and blast-traumatic brain injury 
Neurobiology of disease  2013;54:1-11.
Warfare has long been associated with traumatic brain injury (TBI) in militarized zones. Common forms of TBI can be caused by a physical insult to the head-brain or by the effects of a high velocity blast shock wave generated by the detonation of an explosive device. While both forms of trauma are distinctly different regarding the mechanism of trauma induction, there are striking similarities in the cognitive and emotional status of survivors. Presently, proven effective therapeutics for the treatment of either form of TBI are unavailable. To be able to develop efficacious therapies, studies involving animal models of physical- and blast-TBI are required to identify possible novel or existing medicines that may be of value in the management of clinical events. We examined indices of cognition and anxiety-like behavior and the hippocampal gene transcriptome of mice subjected to both forms of TBI. We identified common behavioral deficits and gene expression regulations, in addition to unique injury-specific forms of gene regulation. Molecular pathways presented a pattern similar to that seen in gene expression. Interestingly, pathways connected to Alzheimer’s disease displayed a markedly different form of regulation depending on the type of TBI. While these data highlight similarities in behavioral outcomes after trauma, the divergence in hippocampal transcriptome observed between models suggests that, at the molecular level, the TBIs are quite different. These models may provide tools to help define therapeutic approaches for the treatment of physical- and blast-TBIs. Based upon observations of increasing numbers of personnel displaying TBI related emotional and behavioral changes in militarized zones, the development of efficacious therapies will become a national if not a global priority.
doi:10.1016/j.nbd.2013.02.006
PMCID: PMC3628969  PMID: 23454194
Physical-traumatic brain injury; Blast-traumatic brain injury; Cognitive dysfunction; Gene expression; Molecular pathway(s); Neurodegeneration; Stem cells; Alzheimer’s disease
2.  Molecular characterization of the transition to mid-life in Caenorhabditis elegans 
Age  2012;35(3):689-703.
We present an initial molecular characterization of a morphological transition between two early aging states. In previous work, an age score reflecting physiological age was developed using a machine classifier trained on images of worm populations at fixed chronological ages throughout their lifespan. The distribution of age scores identified three stable post-developmental states and transitions. The first transition occurs at day 5 post-hatching, where a significant percentage of the population exists in both state I and state II. The temperature dependence of the timing of this transition (Q10 ~ 1.17) is too low to be explained by a stepwise process with an enzymatic or chemical rate-limiting step, potentially implicating a more complex mechanism. Individual animals at day 5 were sorted into state I and state II groups using the machine classifier and analyzed by microarray expression profiling. Despite being isogenic, grown for the same amount of time, and indistinguishable by eye, these two morphological states were confirmed to be molecularly distinct by hierarchical clustering and principal component analysis of the microarray results. These molecular differences suggest that pharynx morphology reflects the aging state of the whole organism. Our expression profiling yielded a gene set that showed significant overlap with those from three previous age-related studies and identified several genes not previously implicated in aging. A highly represented group of genes unique to this study is involved in targeted ubiquitin-mediated proteolysis, including Skp1-related (SKR), F-box-containing, and BTB motif adaptors.
Electronic supplementary material
The online version of this article (doi:10.1007/s11357-012-9401-2) contains supplementary material, which is available to authorized users.
doi:10.1007/s11357-012-9401-2
PMCID: PMC3636400  PMID: 22610697
Machine classifier; Biomarker of aging; Metastable aging state; Microarray analysis
3.  The SIRT1 activator SRT1720 extends lifespan and improves health of mice fed a standard diet 
Cell reports  2014;6(5):836-843.
The prevention or delay of the onset of age-related diseases prolongs survival and improves quality of life while reducing the burden on the health care system. Activation of sirtuin 1 (SIRT1), an NAD+ deacetylase, improves metabolism and confers protection against physiological and cognitive disturbances in old age. SRT1720 is a specific SIRT1 activator that has health and lifespan benefits in adult mice fed a high-fat diet. We found extension in lifespan, delayed onset of age-related metabolic diseases, and improved general health in mice fed a standard diet after SRT1720 supplementation. Inhibition of pro-inflammatory gene expression both in the liver and muscle of SRT1720-treated animals was noted. SRT1720 lowered phosphorylation of NF-κB pathway regulators in vitro only when SIRT1 was functionally present. Combined with our previous work, the current study further supports the beneficial effects of SRT1720 on health across the lifespan in mice.
doi:10.1016/j.celrep.2014.01.031
PMCID: PMC4010117  PMID: 24582957
SRT1720; healthspan; standard diet; mice; longevity; SIRT1
4.  Metformin improves healthspan and lifespan in mice 
Nature communications  2013;4:2192.
Metformin is a drug commonly prescribed to treat patients with type 2 diabetes. Here we show that long-term treatment with metformin (0.1% w/w in diet) starting at middle age extends healthspan and lifespan in male mice, while a higher dose (1% w/w) was toxic. Treatment with metformin mimics some of the benefits of calorie restriction, such as improved physical performance, increased insulin sensitivity, and reduced LDL and cholesterol levels without a decrease in caloric intake. At a molecular level, metformin increases AMP-activated protein kinase activity and increases antioxidant protection, resulting in reductions in both oxidative damage accumulation and chronic inflammation. Our results indicate that these actions may contribute to the beneficial effects of metformin on healthspan and lifespan. These findings are in agreement with current epidemiological data and raise the possibility of metformin-based interventions to promote healthy aging.
doi:10.1038/ncomms3192
PMCID: PMC3736576  PMID: 23900241
5.  Down-Regulation of eIF4GII by miR-520c-3p Represses Diffuse Large B Cell Lymphoma Development 
PLoS Genetics  2014;10(1):e1004105.
Deregulation of the translational machinery is emerging as a critical contributor to cancer development. The contribution of microRNAs in translational gene control has been established however; the role of microRNAs in disrupting the cap-dependent translation regulation complex has not been previously described. Here, we established that elevated miR-520c-3p represses global translation, cell proliferation and initiates premature senescence in HeLa and DLBCL cells. Moreover, we demonstrate that miR-520c-3p directly targets translation initiation factor, eIF4GII mRNA and negatively regulates eIF4GII protein synthesis. miR-520c-3p overexpression diminishes cells colony formation and reduces tumor growth in a human xenograft mouse model. Consequently, downregulation of eIF4GII by siRNA decreases translation, cell proliferation and ability to form colonies, as well as induces cellular senescence. In vitro and in vivo findings were further validated in patient samples; DLBCL primary cells demonstrated low miR-520c-3p levels with reciprocally up-regulated eIF4GII protein expression. Our results provide evidence that the tumor suppressor effect of miR-520c-3p is mediated through repression of translation while inducing senescence and that eIF4GII is a key effector of this anti-tumor activity.
Author Summary
Control of gene expression on the translational level is critical for proper function of major cellular processes and deregulation of translation can promote cellular transformation. Emerging actors in this post-transcriptional gene regulation are small non-coding RNAs referred to as microRNAs (miRNAs). We established that miR-520c-3p represses tumor growth through the repression of eIF4GII, a major structural component of the translation initiation complex. Since translation of most cellular mRNAs is primarily regulated at the level of initiation, this node is becoming a potential target for therapeutic intervention. Identified in this study, tumor suppressor function of miR-520c-3p is mediated through the inhibition of translational factor eIF4GII, resulting in the repression of global translational machinery and induction of senescence in tumor cells. While aging and senescence has been shown to be associated with reduced translation the linkage between translational deregulation and senescence in malignant cells has not been previously described. Lending further clinical significance to our findings, we were able to demonstrate that primary DLBCL samples had elevated levels of eIF4GII while having reciprocally low miR-520c-3p expression.
doi:10.1371/journal.pgen.1004105
PMCID: PMC3907297  PMID: 24497838
6.  Mice Fed Rapamycin Have an Increase in Lifespan Associated with Major Changes in the Liver Transcriptome 
PLoS ONE  2014;9(1):e83988.
Rapamycin was found to increase (11% to 16%) the lifespan of male and female C57BL/6J mice most likely by reducing the increase in the hazard for mortality (i.e., the rate of aging) term in the Gompertz mortality analysis. To identify the pathways that could be responsible for rapamycin's longevity effect, we analyzed the transcriptome of liver from 25-month-old male and female mice fed rapamycin starting at 4 months of age. Few changes (<300 transcripts) were observed in transcriptome of rapamycin-fed males; however, a large number of transcripts (>4,500) changed significantly in females. Using multidimensional scaling and heatmap analyses, the male mice fed rapamycin were found to segregate into two groups: one group that is almost identical to control males (Rapa-1) and a second group (Rapa-2) that shows a change in gene expression (>4,000 transcripts) with more than 60% of the genes shared with female mice fed Rapa. Using ingenuity pathway analysis, 13 pathways were significantly altered in both Rapa-2 males and rapamycin-fed females with mitochondrial function as the most significantly changed pathway. Our findings show that rapamycin has a major effect on the transcriptome and point to several pathways that would likely impact the longevity.
doi:10.1371/journal.pone.0083988
PMCID: PMC3883653  PMID: 24409289
7.  Interrogation of brain miRNA and mRNA expression profiles reveals a molecular regulatory network that is perturbed by mutant huntingtin 
Journal of neurochemistry  2012;123(4):477-490.
Emerging evidence indicates that microRNAs (miRNAs) may play an important role in the pathogenesis of Huntington’s disease (HD). To identify the individual miRNAs that are altered in HD and may therefore regulate a gene network underlying mutant huntingtin-induced neuronal dysfunction in HD, we performed miRNA array analysis combined with mRNA profiling in the cerebral cortex from N171-82Q HD mice. Expression profiles of miRNAs as well as mRNAs in HD mouse cerebral cortex were analyzed and confirmed at different stages of disease progression; the most significant changes of miRNAs in the cerebral cortex were also detected in the striatum of HD mice. Our results revealed a significant alteration of miR-200 family members, miR-200a and miR-200c in the cerebral cortex and the striatum, at the early stage of disease progression in N171-82Q HD mice. We used a coordinated approach to integrate miRNA and mRNA profiling, and applied bioinformatics to predict a target gene network potentially regulated by these significantly altered miRNAs that might be involved in HD disease progression. Interestingly, miR-200a and miR-200c are predicted to target genes regulating synaptic function, neurodevelopment and neuronal survival. Our results suggest that altered expression of miR-200a and miR-200c may interrupt the production of proteins involved in neuronal plasticity and survival, and further investigation of the involvement of perturbed miRNA expression in HD pathogenesis is warranted, and may lead to reveal novel approaches for HD therapy.
doi:10.1111/j.1471-4159.2012.07925.x
PMCID: PMC3472040  PMID: 22906125
miRNA array; Huntington’s disease; gene array; miR-200; Trim2
8.  Response of microchip solid-state laser to external frequency-shifted feedback and its applications 
Scientific Reports  2013;3:2912.
The response of the microchip solid-state Nd:YAG laser, which is subjected to external frequency-shifted feedback, is experimentally and theoretically analysed. The continuous weak response of the laser to the phase and amplitude of the feedback light is achieved by controlling the feedback power level, and this system can be used to achieve contact-free measurement of displacement, vibration, liquid evaporation and thermal expansion with nanometre accuracy in common room conditions without precise environmental control. Furthermore, a strong response, including chaotic harmonic and parametric oscillation, is observed, and the spectrum of this response, as examined by a frequency-stabilised Nd:YAG laser, indicates laser spectral linewidth broadening.
doi:10.1038/srep02912
PMCID: PMC3793223  PMID: 24105389
9.  Age-related changes in microRNA levels in serum 
Aging (Albany NY)  2013;5(10):725-740.
microRNAs (miRNAs) are small noncoding RNAs that post-transcriptionally regulate gene expression by targeting specific mRNAs. Altered expression of circulating miRNAs have been associated with age-related diseases including cancer and cardiovascular disease. Although we and others have found an age-dependent decrease in miRNA expression in peripheral blood mononuclear cells (PBMCs), little is known about the role of circulating miRNAs in human aging. Here, we examined miRNA expression in human serum from young (mean age 30 years) and old (mean age 64 years) individuals using next generation sequencing technology and real-time quantitative PCR. Of the miRNAs that we found to be present in serum, three were significantly decreased in 20 older individuals compared to 20 younger individuals: miR-151a-5p, miR-181a-5p and miR-1248. Consistent with our data in humans, these miRNAs are also present at lower levels in the serum of elderly rhesus monkeys. In humans, miR-1248 was found to regulate the expression of mRNAs involved in inflammatory pathways and miR-181a was found to correlate negatively with the pro-inflammatory cytokines IL-6 and TNFα and to correlate positively with the anti-inflammatory cytokines TGFβ and IL-10. These results suggest that circulating miRNAs may be a biological marker of aging and could also be important for regulating longevity. Identification of stable miRNA biomarkers in serum could have great potential as a noninvasive diagnostic tool as well as enhance our understanding of physiological changes that occur with age.
PMCID: PMC3838776  PMID: 24088671
circulating; miRNA; noncoding RNA; age; aging; biomarker; exRNA; extracellular RNA
10.  Genome-wide modeling of complex phenotypes in Caenorhabditis elegans and Drosophila melanogaster 
BMC Genomics  2013;14:580.
Background
The genetic and molecular basis for many intermediate and end stage phenotypes in model systems such as C. elegans and D. melanogaster has long been known to involve pleiotropic effects and complex multigenic interactions. Gene sets are groups of genes that contribute to multiple biological or molecular phenomena. They have been used in the analysis of large molecular datasets such as microarray data, Next Generation sequencing, and other genomic datasets to reveal pleiotropic and multigenic contributions to phenotypic outcomes. Many model systems lack species specific organized phenotype based gene sets to enable high throughput analysis of large molecular datasets.
Results and discussion
Here, we describe two novel collections of gene sets in C. elegans and D. melanogaster that are based exclusively on genetically determined phenotypes and use a controlled phenotypic ontology. We use these collections to build genome-wide models of thousands of defined phenotypes in both model species. In addition, we demonstrate the utility of these gene sets in systems analysis and in analysis of gene expression-based molecular datasets and show how they are useful in analysis of genomic datasets connecting multigenic gene inputs to complex phenotypes.
Conclusions
Phenotypic based gene sets in both C. elegans and D. melanogaster are developed, characterized, and shown to be useful in the analysis of large scale species-specific genomic datasets. These phenotypic gene set collections will contribute to the understanding of complex phenotypic outcomes in these model systems.
doi:10.1186/1471-2164-14-580
PMCID: PMC3849582  PMID: 23984798
C. elegans; D. melanogaster; Worm; Fly; Aging; Gene set; Phenotype; Ontology; Network; Gene expression
11.  Estimate of nocturnal blood pressure and detection of non-dippers based on clinical or ambulatory monitoring in the inpatient setting 
Background
Ambulatory blood pressure monitoring is regarded as the gold standard for monitoring nocturnal blood pressure (NBP) and is usually performed out of office. Currently, a novel method for monitoring NBP is indispensible in the inpatient setting. The widely used manual BP monitoring procedure has the potential to monitor NBP in the hospital setting. The feasibility and accuracy of manual sphygmomanometer to monitor NBP has not been explored widely.
Methods
A cross-sectional study was conducted at the cardiology department of a university-affiliated hospital to study patients with mild-to-moderate essential hypertension. One hundred and fifty-five patients were recruited to compare BP derived from a manual device and ambulatory BP monitoring (ABPM). The manual BP measurement was performed six times at 22:00, 02:00, 06:00, 10:00, 14:00 and 18:00 h. The measurements at 22:00, 02:00 and 06:00 h were defined as night-time and the others as daytime. ABPM was programmed to measure at 30-min intervals between measurements.
Results
All-day, daytime and night-time BP did not differ significantly from 24-h ambulatory systolic BP [all-day mean difference −0.52±4.67 mmHg, 95% confidence interval (CI) –1.26 to 0.22, P=0.168; daytime mean difference 0.24±5.45 mmHg, 95% CI −0.62 to 1.11, P=0.580; night-time mean difference 0.30±7.22 mmHg, 95% CI −0.84 to 1.45, P=0.601) rather than diastolic BP. There was a strong correlation between clinical and ambulatory BP for both systolic and diastolic BP. On the basis of ABPM, 101 (65%) patients were classified as non-dippers, compared with 106 (68%) by manual sphygmomanometer (P<0.001).
Conclusions
Traditional manual sphygmomanometer provides similar daytime and night-time systolic BP measurements in hospital. Moreover, the detection of non-dippers by manual methods is in good agreement with 24-h ABPM. Further studies are required to confirm the clinical relevance of these findings by comparing the association of NBP in the hospital ward assessed by manual monitoring with preclinical organ damage and cardiovascular and cerebrovascular outcomes.
doi:10.1186/1471-2261-13-37
PMCID: PMC3665662  PMID: 23692688
Nocturnal blood pressure; Ambulatory blood pressure monitoring; Hypertension; Non-dipper
12.  Platelets contribute to the pathogenesis of experimental autoimmune encephalomyelitis 
Circulation Research  2012;110(9):1202-1210.
Rationale
Multiple sclerosis (MS) and its mouse model, experimental autoimmune encephalomyelitis (EAE), are inflammatory disorders of the central nervous system (CNS). The function of platelets in inflammatory and autoimmune pathologies is thus far poorly defined.
Objective
Here we addressed the role of platelets in mediating CNS inflammation in EAE.
Results
We found that platelets were present in human MS lesions as well as in the CNS of mice subjected to EAE but not in the CNS from control non-diseased mice. Platelet depletion at the effector-inflammatory phase of EAE in mice resulted in significantly ameliorated disease development and progression. EAE suppression upon platelet depletion was associated with reduced recruitment of leukocytes to the inflamed CNS, as assessed by intravital microscopy, and with a blunted inflammatory response. The platelet-specific receptor glycoprotein Ib alpha (GPIbα) promotes both platelet adhesion as well as inflammatory actions of platelets, and, targeting of GPIbα attenuated EAE in mice. Moreover, targeting another platelet adhesion receptor, glycoprotein IIb/IIIa (GPIIb/IIIa) also reduced EAE severity in mice.
Conclusions
Thus, platelets contribute to the pathogenesis of EAE by promoting CNS inflammation. Targeting platelets may therefore represent an important new therapeutic approach for MS treatment.
doi:10.1161/CIRCRESAHA.111.256370
PMCID: PMC3382058  PMID: 22456181
Platelets; EAE; inflammation; autoimmune disease
13.  Non-Steroidal Anti-inflammatory Drugs Decrease E2F1 Expression and Inhibit Cell Growth in Ovarian Cancer Cells 
PLoS ONE  2013;8(4):e61836.
Epidemiological studies have shown that the regular use of non-steroidal anti-inflammatory (NSAIDs) drugs is associated with a reduced risk of various cancers. In addition, in vitro and experiments in mouse models have demonstrated that NSAIDs decrease tumor initiation and/or progression of several cancers. However, there are limited preclinical studies investigating the effects of NSAIDs in ovarian cancer. Here, we have studied the effects of two NSAIDs, diclofenac and indomethacin, in ovarian cancer cell lines and in a xenograft mouse model. Diclofenac and indomethacin treatment decreased cell growth by inducing cell cycle arrest and apoptosis. In addition, diclofenac and indomethacin reduced tumor volume in a xenograft model of ovarian cancer. To identify possible molecular pathways mediating the effects of NSAID treatment in ovarian cancer, we performed microarray analysis of ovarian cancer cells treated with indomethacin or diclofenac. Interestingly, several of the genes found downregulated following diclofenac or indomethacin treatment are transcriptional target genes of E2F1. E2F1 was downregulated at the mRNA and protein level upon treatment with diclofenac and indomethacin, and overexpression of E2F1 rescued cells from the growth inhibitory effects of diclofenac and indomethacin. In conclusion, NSAIDs diclofenac and indomethacin exert an anti-proliferative effect in ovarian cancer in vitro and in vivo and the effects of NSAIDs may be mediated, in part, by downregulation of E2F1.
doi:10.1371/journal.pone.0061836
PMCID: PMC3634839  PMID: 23637916
14.  Single-round of antigen receptor signaling programs naïve B cells to receive T cell help 
Immunity  2010;32(3):355-366.
SUMMARY
To simulate transient B cell activation that is the likely initiator of T-dependent responses, we examined the molecular and functional consequences of a single-round of immunoglobulin M (IgM) signaling. This form of activation triggered early cytosolic signaling and the transcription factor NF-κB activation indistinguishably from conventional continuous IgM cross-linking, but did not induce G1 progression. However, single-round IgM signaling changed the expression of chemokine and chemokine receptor genes implicated in initiating T-dependent responses, as well as accentuated responsiveness to CD40 signaling. Several features of single-round IgM signaling in vitro were recapitulated in B cells after short-term exposure to antigen in vivo. We propose that transient BCR signals prime B cells to receive T cell help by increasing the probability of B-T encounter and creating a cellular environment that is hyper-responsive to CD40 signaling.
doi:10.1016/j.immuni.2010.02.013
PMCID: PMC3607434  PMID: 20226693
15.  Long Noncoding RNA MALAT1 Controls Cell Cycle Progression by Regulating the Expression of Oncogenic Transcription Factor B-MYB 
PLoS Genetics  2013;9(3):e1003368.
The long noncoding MALAT1 RNA is upregulated in cancer tissues and its elevated expression is associated with hyper-proliferation, but the underlying mechanism is poorly understood. We demonstrate that MALAT1 levels are regulated during normal cell cycle progression. Genome-wide transcriptome analyses in normal human diploid fibroblasts reveal that MALAT1 modulates the expression of cell cycle genes and is required for G1/S and mitotic progression. Depletion of MALAT1 leads to activation of p53 and its target genes. The cell cycle defects observed in MALAT1-depleted cells are sensitive to p53 levels, indicating that p53 is a major downstream mediator of MALAT1 activity. Furthermore, MALAT1-depleted cells display reduced expression of B-MYB (Mybl2), an oncogenic transcription factor involved in G2/M progression, due to altered binding of splicing factors on B-MYB pre-mRNA and aberrant alternative splicing. In human cells, MALAT1 promotes cellular proliferation by modulating the expression and/or pre-mRNA processing of cell cycle–regulated transcription factors. These findings provide mechanistic insights on the role of MALAT1 in regulating cellular proliferation.
Author Summary
The mammalian genome encodes large number of long non protein-coding RNAs (lncRNAs). These lncRNAs are suggested to regulate key biological processes (including cellular proliferation and differentiation), and aberrant expression of these is associated with cancer. However, only a few of these lncRNAs have been functionally validated in biological or disease processes. MALAT1, an abundant nuclear-retained lncRNA, is overexpressed in several cancers, and its elevated expression has been associated with hyper-proliferation and metastasis. However, the underlying mechanism behind this deregulation and its association with cancer is poorly understood. Here, we establish the role of MALAT1 in the cell cycle pathway and propose the molecular mechanism of its function during normal cell cycle progression. MALAT1 RNA levels are differentially regulated and critical for normal cell cycle progression. Depletion of MALAT1 results in cell cycle arrest with significantly reduced cellular proliferation, simultaneously leading to activation of p53 and its target genes. Further, the accurate levels of MALAT1 in the cell are extremely crucial for expression and activity of the oncogenic transcription factor B-MYB, which is involved in G2/M progression. Our data indicates that the cancer-associated MALAT1 RNA regulates cellular proliferation by modulating the expression and/or pre-mRNA processing of cell cycle–regulated transcription factors.
doi:10.1371/journal.pgen.1003368
PMCID: PMC3605280  PMID: 23555285
17.  FACS purification of immunolabeled cell types from adult rat brain 
Journal of neuroscience methods  2011;203(1):10-18.
Molecular analysis of brain tissue is greatly complicated by having many different classes of neurons and glia interspersed throughout the brain. Fluorescence-activated cell sorting (FACS) has been used to purify selected cell types from brain tissue. However, its use has been limited to brain tissue from embryos or transgenic mice with promoter-driven reporter genes. To overcome these limitations, we developed a FACS procedure for dissociating intact cell bodies from adult wild-type rat brains and sorting them using commercially available antibodies against intracellular and extracellular proteins. As an example, we isolated neurons using a NeuN antibody and confirmed their identity using microarray and real time PCR of mRNA from the sorted cells. Our FACS procedure allows rapid, high-throughput, quantitative assays of molecular alterations in identified cell types with widespread applications in neuroscience.
doi:10.1016/j.jneumeth.2011.08.045
PMCID: PMC3221768  PMID: 21911005
glia; genes; microarray; qPCR
18.  Molecular changes in brain aging and Alzheimer’s disease are mirrored in experimentally silenced cortical neuron networks 
Neurobiology of aging  2010;33(1):205.e1-205.e18.
Activity-dependent modulation of neuronal gene expression promotes neuronal survival and plasticity, and neuronal network activity is perturbed in aging and Alzheimer’s disease (AD). Here we show that cerebral cortical neurons respond to chronic suppression of excitability by downregulating the expression of genes and their encoded proteins involved in inhibitory transmission (GABAergic and somatostatin) and Ca2+ signaling; alterations in pathways involved in lipid metabolism and energy management are also features of silenced neuronal networks. A molecular fingerprint strikingly similar to that of diminished network activity occurs in the human brain during aging and in AD, and opposite changes occur in response to activation of N-methyl-D-aspartate (NMDA) and brain-derived neurotrophic factor (BDNF) receptors in cultured cortical neurons and in mice in response to an enriched environment or electroconvulsive shock. Our findings suggest that reduced inhibitory neurotransmission during aging and in AD may be the result of compensatory responses that, paradoxically, render the neurons vulnerable to Ca2+-mediated degeneration.
doi:10.1016/j.neurobiolaging.2010.08.012
PMCID: PMC3027841  PMID: 20947216
Alzheimer’s disease; Aging; GABA; Activity; Homeostatic disinhibition; Interneuron; Calcium; Synaptic scaling
19.  Toll-like receptor 2 (TLR2)-TLR9 crosstalk dictates IL-12 family cytokine production in microglia 
Glia  2011;60(1):29-42.
Microglia are the resident mononuclear phagocytes of the CNS parenchyma and represent an initial line of defense against invading microorganisms. Microglia utilize Toll-like receptors (TLRs) for pathogen recognition and TLR2 specifically senses conserved motifs of Gram-positive bacteria including lipoproteins, lipoteichoic acids, and peptidoglycan (PGN) leading to cytokine/chemokine production. Interestingly, primary microglia derived from TLR2 knockout (KO) mice over-expressed numerous IL-12 family members, including IL-12p40, IL-12p70, and IL-27 in response to intact S. aureus, but not the less structurally complex TLR2 ligands Pam3CSK4 or PGN. The ability of intact bacteria to augment IL-12 family member expression was specific for Gram-positive organisms since numerous Gram-negative strains were unable to elicit exaggerated responses in TLR2 KO microglia. Inhibition of SYK or IRAK4 signaling did not impact heightened IL-12 family member production in S. aureus-treated TLR2 KO microglia, whereas PI3K, MAPK, and JNK inhibitors were all capable of restoring exaggerated cytokine expression to WT levels. Additionally, elevated IL-12 production in TLR2 KO microglia was ablated by a TLR9 antagonist, suggesting that TLR9 drives IL-12 family member production following exposure to intact bacteria that remains unchecked in the absence of TLR2 signaling. Collectively, these findings indicate crosstalk between TLR2 and TLR9 pathways to regulate IL-12 family member production by microglia. The summation of TLR signals must be tightly controlled to ensure the timely cessation and/or fine tuning of cytokine signaling to avoid non-specific bystander damage due to sustained IL-12 release.
doi:10.1002/glia.21243
PMCID: PMC3217087  PMID: 21901759
S. aureus; TLR2; TLR9; microglia; MAPK; JNK; PI3K
20.  Age-associated alterations in inducible gene transcription in human CD4+ T lymphocytes 
Aging (Albany NY)  2013;5(1):18-36.
Age associated immune dysregulation results in a pro-inflammatory state and increased susceptibility to infections and autoimmune diseases. Studies show that signaling initiated at the T cell antigen receptor (TCR) is impaired in CD4+ T cells from old compared to young mice. Here we examined TCR-inducible gene expression changes in CD4+ T cells during human aging. We reveal a dichotomy in gene expression mediated by the inducible transcription factor NF-κB. Most NF-κB target genes are not induced in a sustained manner in cells derived from older compared to younger individuals. However, a subset of NF-κB target genes including genes associated with chronic pro-inflammatory state in the elderly, such as interleukin 1 and 6, continue to be up-regulated even in the absence of NF-κB induction. In addition, we identify other widespread changes in gene expression between cells derived from older and younger individuals. Surprisingly, many of the most noteworthy age-associated changes in human CD4+ T cells differ from those seen in murine models. Our studies provide the first view of age-associated alteration of TCR-inducible gene expression in human CD4+ T cells.
PMCID: PMC3616229  PMID: 23385138
human; CD4+ T cell; NF-κB; aging; aene expression
21.  Aging-kb: a knowledge base for the study of the Aging process 
Mechanisms of Ageing and Development  2011;132(11-12):592-594.
As the science of the aging process moves forward, a recurring challenge is the integration of multiple types of data and information with classical aging theory while disseminating that information to the scientific community. Here we present AGING-kb, a public knowledge base with the goal of conceptualizing and presenting fundamental aspects of the study of the aging process. Aging-kb has two interconnected parts, the Aging-kb tree and the Aging Wiki. The Aging-kb tree is a simple intuitive dynamic tree hierarchy of terms describing the field of aging from the general to the specific. This enables the user to see relationships between areas of aging research in a logical comparative fashion. The second part is a specialized Aging Wiki which allows expert definition, description, supporting information, and documentation of each aging keyword term found in the Aging-kb tree. The Aging Wiki allows community participation in describing and defining concepts and terms in the Wiki format. This aging knowledge base provides a simple intuitive interface to the complexities of aging.
http://aging-kb.nia.nih.gov
doi:10.1016/j.mad.2011.10.008
PMCID: PMC3287063  PMID: 22100666
22.  Identification and characterization of unique tumoricidal genes in rat umbilical cord matrix stem cells 
Molecular pharmaceutics  2011;8(5):1549-1558.
Rat umbilical cord matrix stem cells (UCMSC) have been shown to exhibit a remarkable ability to control rat mammary adenocarcinoma (Mat B III) cell proliferation both in vivo and in vitro. To study the underlying mechanisms and genes involved in Mat B III growth attenuation, total RNA was extracted from the naïve rat UCMSC alone and those co-cultured with Mat B III in Transwell culture dishes. Gene expression profiles of naive rat UCMSC alone and those co-cultured with Mat B III cells were investigated by microarray analysis using an Illumina RatRef-12 Expression BeadChip. The comparison of gene expression profiles between untreated and co-cultured rat UCMSC identified five up-regulated candidate genes (follistatin (FST), sulfatase1 (SULF-1), glucose phosphate isomerase (GPI), HtrA serine peptidase (HTRA1), and adipocyte differentiation-related protein (ADRP)) and two down-regulated candidate genes (transforming growth factor, beta-induced, 68kDa (TGFβI) and podoplanin (PDPN)) based upon the following screening criteria: 1) expression of the candidate genes should show at least a 1.5 fold change in rat UCMSC co-cultured with Mat B III cells; 2) candidate genes encode secretory proteins; and 3) they encode cell growth-related proteins. Following confirmation of gene expression by real time-PCR, ADRP, SULF-1 and GPI were selected for further analysis. Addition of specific neutralizing antibodies against these three gene products individually in co-cultures of 1:20 rat UCMSC:Mat B III cells significantly increased cell proliferation, implying that these gene products are produced under the co-cultured condition and functionally attenuate cell growth. Immunoprecipitation followed by Western blot analysis demonstrated that these proteins are indeed secreted into the culture medium. Individual over-expression of these three genes in rat UCMSC significantly enhanced UCMSC-dependent inhibition of cell proliferation in co-culture. These results suggest that ADRP, SULF-1 and GPI act as tumor suppressor genes, and these genes might be involved in rat UCMSC-dependent growth attenuation of rat mammary tumors.
doi:10.1021/mp2001582
PMCID: PMC3202613  PMID: 21851062
Rat umbilical cord matrix stem cells; Rat mammary tumor cells; Mat B III; Microarray; Real time PCR; Thymidine uptake; Tumor suppressor genes; ADRP; GPI; SULF-1
23.  Metabolic Context Regulates Distinct Hypothalamic Transcriptional Responses to Antiaging Interventions 
The hypothalamus is an essential relay in the neural circuitry underlying energy metabolism that needs to continually adapt to changes in the energetic environment. The neuroendocrine control of food intake and energy expenditure is associated with, and likely dependent upon, hypothalamic plasticity. Severe disturbances in energy metabolism, such as those that occur in obesity, are therefore likely to be associated with disruption of hypothalamic transcriptomic plasticity. In this paper, we investigated the effects of two well-characterized antiaging interventions, caloric restriction and voluntary wheel running, in two distinct physiological paradigms, that is, diabetic (db/db) and nondiabetic wild-type (C57/Bl/6) animals to investigate the contextual sensitivity of hypothalamic transcriptomic responses. We found that, both quantitatively and qualitatively, caloric restriction and physical exercise were associated with distinct transcriptional signatures that differed significantly between diabetic and non-diabetic mice. This suggests that challenges to metabolic homeostasis regulate distinct hypothalamic gene sets in diabetic and non-diabetic animals. A greater understanding of how genetic background contributes to hypothalamic response mechanisms could pave the way for the development of more nuanced therapeutics for the treatment of metabolic disorders that occur in diverse physiological backgrounds.
doi:10.1155/2012/732975
PMCID: PMC3427989  PMID: 22934110
24.  IL-10 transcription is negatively regulated by BAF180, a component of the SWI/SNF chromatin remodeling enzyme 
BMC Immunology  2012;13:9.
Background
SWI/SNF chromatin remodeling enzymes play a critical role in the development of T helper lymphocytes, including Th2 cells, and directly program chromatin structure at Th2 cytokine genes. Different versions of SWI/SNF complexes, including BAF and PBAF, have been described based on unique subunit composition. However, the relative role of BAF and PBAF in Th cell function and cytokine expression has not been reported.
Results
Here we examine the role of the PBAF SWI/SNF complex in Th cell development and gene expression using mice deficient for a PBAF-specific component, BAF180. We find that T cell development in the thymus and lymphoid periphery is largely normal when the BAF180 gene is deleted late in thymic development. However, BAF180-deficient Th2 cells express high levels of the immunoregulatory cytokine IL-10. BAF180 binds directly to regulatory elements in the Il-10 locus but is replaced by BAF250 BAF complexes in the absence of BAF180, resulting in increased histone acetylation and CBP recruitment to the IL-10 locus.
Conclusions
These results demonstrate that BAF180 is a repressor of IL-10 transcription in Th2 cells and suggest that the differential recruitment of different SWI/SNF subtypes can have direct consequences on chromatin structure and gene transcription.
doi:10.1186/1471-2172-13-9
PMCID: PMC3313858  PMID: 22336179
25.  Claudin-7 Is Frequently Overexpressed in Ovarian Cancer and Promotes Invasion 
PLoS ONE  2011;6(7):e22119.
Background
Claudins are tight junction proteins that are involved in tight junction formation and function. Previous studies have shown that claudin-7 is frequently upregulated in epithelial ovarian cancer (EOC) along with claudin-3 and claudin-4. Here, we investigate in detail the expression patterns of claudin-7, as well as its possible functions in EOC.
Methodology/Principal Findings
A total of 95 ovarian tissue samples (7 normal ovarian tissues, 65 serous carcinomas, 11 clear cell carcinomas, 8 endometrioid carcinomas and 4 mucinous carcinomas) were studied for claudin-7 expression. In real-time RT-PCR analysis, the gene for claudin-7, CLDN7, was found to be upregulated in all the tumor tissue samples studied. Similarly, immunohistochemical analysis and western blotting showed that claudin-7 protein was significantly overexpressed in the vast majority of EOCs. Small interfering RNA-mediated knockdown of claudin-7 in ovarian cancer cells led to significant changes in gene expression as measured by microarrays and validated by RT-PCR and immunoblotting. Analyses of the genes differentially expressed revealed that the genes altered in response to claudin-7 knockdown were associated with pathways implicated in various molecular and cellular functions such as cell cycle, cellular growth and proliferation, cell death, development, and cell movement. Through functional experiments in vitro, we found that both migration and invasion were altered in cells where CLDN7 had been knocked down or overexpressed. Interestingly, claudin-7 expression was associated with a net increase in invasion, but also with a decrease in migration.
Conclusion/Significance
Our work shows that claudin-7 is significantly upregulated in EOC and that it may be functionally involved in ovarian carcinoma invasion. CLDN7 may therefore represent potential marker for ovarian cancer detection and a target for therapy.
doi:10.1371/journal.pone.0022119
PMCID: PMC3137611  PMID: 21789222

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