Expression In Central Nervous System Research Articles

The CNS glycoprotein Shadoo has PrPC-like protective properties and displays reduced levels in prion infections

The cellular prion protein, PrPC, is neuroprotective in a number of settings and in particular prevents cerebellar degeneration mediated by CNS-expressed Doppel or internally deleted PrP (‘ΔPrP'). This paradigm has facilitated mapping of activity determinants in PrPC and implicated a cryptic PrPC-like protein, ‘π'. Shadoo (Sho) is a hypothetical GPI-anchored protein encoded by the Sprn gene, exhibiting homology and domain organization similar to the N-terminus of PrP. Here we demonstrate Sprn expression and Sho protein in the adult CNS. Sho expression overlaps PrPC, but is low in cerebellar granular neurons (CGNs) containing PrPC and high in PrPC-deficient dendritic processes. In Prnp0/0 CGNs, Sho transgenes were PrPC-like in their ability to counteract neurotoxic effects of either Doppel or ΔPrP. Additionally, prion-infected mice exhibit a dramatic reduction in endogenous Sho protein. Sho is a candidate for π, and since it engenders a PrPC-like neuroprotective activity, compromised neuroprotective activity resulting from reduced levels may exacerbate damage in prion infections. Sho may prove useful in deciphering several unresolved facets of prion biology.

MicroRNAs show a wide diversity of expression profiles in the developing and mature central nervous system

A comprehensive analysis of the neuroanatomical expression profiles of 38 abundant conserved miRNAs in developing and adult zebrafish brain was performed.

Integrating transcription factor binding site information with gene expression datasets

Microarrays are widely used to measure gene expression differences between sets of biological samples. Many of these differences will be due to differences in the activities of transcription factors. In principle, these differences can be detected by associating motifs in promoters with differences in gene expression levels between the groups. In practice, this is hard to do. We combine correspondence analysis, between group analysis and co-inertia analysis to determine which motifs, from a database of promoter motifs, are strongly associated with differences in gene expression levels. Given a database of motifs and gene expression levels from a set of arrays, the method produces a ranked list of motifs associated with any specified split in the arrays. We give an example using the Gene Atlas compendium of gene expression levels for human tissues where we search for motifs that are associated with expression in central nervous system (CNS) or muscle tissues. Most of the motifs that we find are known from previous work to be strongly associated with expression in CNS or muscle. We give a second example using a published prostate cancer dataset where we can simply and clearly find which transcriptional pathways are associated with differences between benign and metastatic samples. The source code is freely available upon request from the authors.

Bone morphogenetic protein‐4 expression in the adult rat brain

Bone morphogenetic protein-4 (BMP4) is a member of the transforming growth factor-beta (TGF-beta) superfamily and plays important roles in multiple biological events. Although BMP4 expression has been well described in the early development of the central nervous system (CNS), little information is available on its expression in the adult CNS. Therefore, we investigated BMP4 expression in the adult rat CNS by using immunohistochemistry. BMP4 is intensely expressed in most neurons and their dendrites. In addition, intense BMP4 expression was also observed in the neuropil of the gray matters where high plasticity is reported, such as the molecular layer of the cerebellum and the superficial layer of the superior colliculus. Furthermore, we found that astrocytes also express BMP4 protein. These data indicate that BMP4 is more widely expressed throughout the adult CNS than previously reported, and its continued abundant expression in the adult brain strongly supports the idea that BMP4 plays pivotal roles also in the adult brain.

A novel RT‐PCR method for the quantitative analysis of microRNA expression: miR‐1/124 expression in mouse CNS

More than 700 microRNAs have been cloned, and the functions of these molecules in developmental timing, cell proliferation, and cancer have been investigated widely. In this report, we describe a novel RT-PCR method for the quantitative analysis of microRNA expression. The use of this method in a model study revealed differential expression of miR-1 and -124 in mouse organs. Specifically, the new method revealed that miR-124 concentrations in the mouse CNS (central nervous system) were more than 100 times those in other organs. By contrast, miR-1 expression in the CNS was 100 – 1,000 times lower than that in skeletal muscle and heart. Moreover, the new method revealed regional differences in miR-124 expression within the CNS: expression ratios versus the cerebral cortex were 60.7% for the cerebellum and 35.4% for the spinal cord. These results suggest that our method would be a powerful new tool for studies of microRNA expression.

A functional screen for sonic hedgehog regulatory elements across a 1 Mb interval identifies long-range ventral forebrain enhancers

The secreted protein sonic hedgehog (Shh) plays an integral role in forming the ventral midline of the vertebrate central nervous system (CNS). In the absence of Shh function, ventral midline development is perturbed resulting in holoprosencephaly (HPE), a structural malformation of the brain, as well as in neuronal patterning and path finding defects along the length of the anteroposterior neuraxis. Central to the understanding of ventral neural tube development is how Shh transcription is regulated in the CNS. To address this issue, we devised an enhancer trap assay to systematically screen 1 Mb of DNA surrounding the Shh locus for the ability to target reporter gene expression to sites of Shh transcription in transgenic mouse embryos. This analysis uncovered six enhancers distributed over 400 kb, the combined activity of which covered all sites of Shh expression in the mouse embryonic CNS from the ventral forebrain to the posterior extent of the spinal cord. To evaluate the relative contribution of these enhancers to the overall pattern of Shh expression, individual elements were deleted in the context of a transgenic Bac reporter assay. Redundant mechanisms were found to control Shh-like reporter activity in the ventral spinal cord, hindbrain and regions of the telencephalon, whereas unique elements regulated Shh-like expression in the ventral midbrain, the majority of the ventral diencephalon and parts of the telencephalon. Three ventral forebrain enhancers locate on the distal side of translocation breakpoints that occurred upstream of Shh in human cases of HPE, suggesting that displacement of these regulatory elements from the Shh promoter is a likely cause of HPE in these individuals.

Effects of Prolonged Treatment With the Opiate Tramadol on Prodynorphin Gene Expression in Rat CNS

A low abuse liability is reported for tramadol, an analgesic drug centrally acting through either opioid or nonopioid mechanisms. In this paper, we evaluated the effects of the repeated administration (7 d) of different doses of tramadol (10, 20, and 80 mg/kg, intraperitoneally) on the opioid precursor prodynorphin biosynthesis, in comparison with morphine (10 mg/kg, intraperitoneally), in the rat central nervous system (CNS). Northern analysis showed that morphine and tramadol produced different effects. While morphine caused a downregulation of prodynorphin mRNA levels in all investigated areas (hypothalamus, hippocampus, and striatum), tramadol did not cause any significant change in the striatum, and did not decrease prodynorphin biosynthesis in the hypothalamus and in the hippocampus, at nontoxic doses (10 and 20 mg/kg). The highest dose of tramadol (80 mg/kg) decreased prodynorphin mRNA levels in the hypothalamus and the hippocampus but not in the striatum. These data give some information on tramadol effects at molecular level in the CNS. They indicate that the alterations of prodynorphin gene expression caused by tramadol and morphine show a different pattern that may be related to the different abuse potential of the two analgesic drugs.

Targeted gene therapy to antigen-presenting cells in the central nervous system using hematopoietic stem cells

Background: Hematopoietic stem cells (HSC) have been previously used as vectors for gene therapy of systemic disease. The effectiveness of HSC-mediated gene therapy largely depends on efficient gene delivery into long-term repopulating progenitors and targeted transgene expression in an appropriate progeny of the transduced pluripotent HSCs. In the present study, we examined the feasibility of using HSC transduced with self-inactivating (SIN) lentiviral vectors for the delivery of gene therapy to the central nervous system (CNS).Material and methods: We constructed two SIN lentiviral vectors, EF.GFP and DR.GFP, to express the green fluorescent protein (GFP) gene controlled solely by the promoter of either a housekeeping gene EF-1α or the human HLA-DRα gene, which is selectively expressed in antigen-presenting cells.Results: We demonstrated that both vectors efficiently transduced human pluripotent CD34+ cells capable of engrafting NOD/SCID mice. Only the DR.GFP vector mediated transgene expression in the murine CNS containing human HLA-DR+ cells. These cells express surface markers characteristic of resident CNS microglia. Furthermore, human dendritic cells derived from transduced and engrafted human cells potently stimulated allogeneic T cell proliferation.Conclusions: The present study demonstrated successful targeting of transgene expression to CNS microglia after stable gene transduction of pluripotent HSC.

Transthyretin knockouts are a new mouse model for increased neuropeptide Y

Transthyretin (TTR) has access to the brain and nerve through the blood and cerebrospinal fluid. To investigate TTR function in nervous system homeostasis, differential gene expression in wild-type (WT) and TTR knockout (KO) mice was assessed. Peptidylglycine alpha-amidating monooxygenase (PAM), the rate-limiting enzyme in neuropeptide maturation, is overexpressed in the peripheral (PNS) and central nervous system (CNS) of TTR KOs that, consequently, display increased neuropeptide Y (NPY) levels. NPY acts on energy homeostasis by increasing white adipose tissue lipoprotein lipase (LPL) and decreasing thermogenesis; accordingly, we show increased LPL expression and activity in white adipose tissue, PNS, and CNS as well as decreased body temperature in TTR KOs. Associated to increased NPY levels, TTR KOs display increased carbohydrate consumption and preference. In neuronal cells, absence of TTR is related to increased PAM activity, NPY levels and LPL expression, reinforcing that TTR is involved in neuropeptide maturation and that increased NPY correlates with LPL overexpression in the nervous system. Furthermore, we provide molecular insights to the reduced depressive behavior of TTR KOs, as NPY is anti-depressant. Our findings demonstrate that TTR KOs are a model for increased NPY and that TTR plays a role in nervous system physiology.

Purkinje neuron physiology is altered by the inflammatory factor interleukin‐6

The cytokine interleukin-6 (IL-6) is produced by cells of the central nervous system (CNS) during a variety of neuroinflammatory states, in which it is thought to play a role in neuroprotection and/or neuropathology associated with neurological disease. In addition, CNS expression of IL-6 during non-pathological conditions may also occur, although the conditions for such IL-6 production remain elusive. Expression of IL-6 and its receptor and signal transducing elements by neurons and glia within the cerebellum implicate a role of IL-6 in modulating cerebellar function under normal conditions and in contributing to pathology and pathophysiology within the cerebellum during CNS disease states. Evidence for such a role of IL-6 comes from studies using transgenic mice that chronically express IL-6 within the CNS. These mice exhibit profound cerebellar pathology and significant alterations of Purkinje neuron electrical and synaptic activity. Additional evidence comes from in vitro studies using primary cultures of cerebellar cortex that have been chronically exposed to exogenously applied IL-6. Consistent with the IL-6 transgenic mice, chronic IL-6 treated Purkinje neurons in culture exhibit alterations of endogenous electrophysiological properties as well as changes in intracellular Ca2+ homeostasis and signaling. Despite these changes in Purkinje neuron physiology, chronic IL-6 does not affect the survival or morphology of Purkinje neurons in culture. Thus, by itself, IL-6 is able to modulate key components of cerebellar circuitry during periods of chronic expression, such as during neuroinflammation, and may be an important player in the movement disorders associated with a number of CNS disease states.

Targeted expression of IGF-1 in the central nervous system fails to protect mice from experimental autoimmune encephalomyelitis

Insulin-like growth factor 1 (IGF-1) has been identified as a critical molecule in the induction of myelination in the central nervous system (CNS). Systemic injection of IGF-1 has been shown to have a varied and transiently protective effect on the clinical course of experimental autoimmune encephalomyelitis (EAE). Since systemic IGF-1 can also modulate peripheral immune lymphocytes, we examined whether a sustained and local delivery of IGF-1 into the spinal cord would have any influence on the chronic course of EAE in C57/BL6 mice. The capability of adeno-associated virus (AAV) to be retrogradely transported efficiently from muscle to motor neurons of the spinal cord was used to overcome the difficulty routinely encountered when attempting chronic delivery of molecules into the CNS. We demonstrate that AAV-mediated delivery of IGF-1 in CNS did not have any beneficial effect on the clinical course of EAE. Injection of AAV-IGF1 after induction of the disease worsened the clinical symptoms. Furthermore, CNS expression of IGF-1 did not affect the pathogenic anti-MOG T cell response, as examined by proliferation and cytokine secretion. Thus, enhanced expression of IGF-1 in the CNS during inflammation does not have a significant effect on myelination. These data have important implications for the potential use of IGF-1 in the treatment of multiple sclerosis.

Chronic expression of monocyte chemoattractant protein‐1 in the central nervous system causes delayed encephalopathy and impaired microglial function in mice

Increased central nervous system (CNS) levels of monocyte chemoattractant protein 1 [CC chemokine ligand 2 (CCL2) in the systematic nomenclature] have been reported in chronic neurological diseases such as human immunodeficiency virus type 1-associated dementia, amyotrophic lateral sclerosis, and multiple sclerosis. However, a pathogenic role for CCL2 has not been confirmed, and there is no established model for the effects of chronic CCL2 expression on resident and recruited CNS cells. We report that aged (>6 months) transgenic (tg) mice expressing CCL2 under the control of the human glial fibrillary acidic protein promoter (huGFAP-CCL2hi tg+ mice) manifested encephalopathy with mild perivascular leukocyte infiltration, impaired blood brain barrier function, and increased CD45-immunoreactive microglia, which had morphologic features of activation. huGFAP-CCL2hi tg+ mice lacking CC chemokine receptor 2 (CCR2) were normal, showing that chemokine action via CCR2 was required. Studies of cortical slice preparations using video confocal microscopy showed that microglia in the CNS of huGFAP-CCL2hi tg+ mice were defective in expressing amoeboid morphology. Treatment with mutant CCL2 peptides, a receptor antagonist and an obligate monomer, also suppressed morphological transformation in this assay, indicating a critical role for CCL2 in microglial activation and suggesting that chronic CCL2 exposure desensitized CCR2 on microglia, which in the CNS of huGFAP-CCL2hi tg+ mice, did not up-regulate cell-surface expression of major histocompatibility complex class II, CD11b, CD11c, or CD40, in contrast to recruited perivascular macrophages that expressed enhanced levels of these markers. These results indicate that huGFAP-CCL2hi tg+ mice provide a useful model to study how chronic CNS expression of CCL2 alters microglial function and CNS physiology.

Comparison of inflammatory and acute-phase responses in the brain and peripheral organs of the ME7 model of prion disease.

Chronic neurodegenerative diseases such as prion disease and Alzheimer's disease (AD) are reported to be associated with microglial activation and increased brain and serum cytokines and acute-phase proteins (APPs). Unlike AD, prion disease is also associated with a peripheral component in that the presumed causative agent, PrPSc, also accumulates in the spleen and other lymphoreticular organs. It is unclear whether the reported systemic acute-phase response represents a systemic inflammatory response to prion disease or merely reflects central nervous system (CNS) inflammation. For this study, we investigated whether intracerebrally initiated prion disease (ME7 model) provokes splenic, hepatic, or brain inflammatory and acute-phase responses. We detected no significant elevation of proinflammatory cytokines or activation of macrophages in the spleens of these animals, despite clear PrPSc deposition. Similarly, at 19 weeks we detected no significant elevation of transcripts for the APPs serum amyloid A, complement C3, pentraxin 3, and alpha2-antiplasmin in the liver, despite CNS neurodegeneration and splenic PrPSc deposition at this time. However, despite the low CNS expression levels of proinflammatory cytokines, there was robust expression of these APPs in degenerating brains. These findings suggest that PrPSc is not a stimulus for splenic macrophages and that neither peripheral PrPSc deposition nor CNS neurodegeneration is sufficient to produce a systemic acute-phase response. We also propose that serum cytokine and APP measurements are not useful during preclinical disease. Possible consequences of the clear chronic elevation of APPs in the CNS are discussed.

Impaired insulin and insulin-like growth factor expression and signaling mechanisms in Alzheimer's disease – is this type 3 diabetes?

The neurodegeneration that occurs in sporadic Alzheimer's disease (AD) is consistently associated with a number of characteristic histopathological, molecular, and biochemical abnormalities, including cell loss, abundant neurofibrillary tangles and dystrophic neurites, amyloid-beta deposits, increased activation of pro-death genes and signaling pathways, impaired energy metabolism/mitochondrial function, and evidence of chronic oxidative stress. The general inability to convincingly link these phenomena has resulted in the emergence and propagation of various heavily debated theories that focus on the role of one particular element in the pathogenesis of all other abnormalities. However, the accumulating evidence that reduced glucose utilization and deficient energy metabolism occur early in the course of disease, suggests a role for impaired insulin signaling in the pathogenesis of AD. The present work demonstrates extensive abnormalities in insulin and insulin-like growth factor type I and II (IGF-I and IGF-II) signaling mechanisms in brains with AD, and shows that while each of the corresponding growth factors is normally made in central nervous system (CNS) neurons, the expression levels are markedly reduced in AD. These abnormalities were associated with reduced levels of insulin receptor substrate (IRS) mRNA, tau mRNA, IRS-associated phosphotidylinositol 3-kinase, and phospho-Akt (activated), and increased glycogen synthase kinase-3beta activity and amyloid precursor protein mRNA expression. The strikingly reduced CNS expression of genes encoding insulin, IGF-I, and IGF-II, as well as the insulin and IGF-I receptors, suggests that AD may represent a neuro-endocrine disorder that resembles, yet is distinct from diabetes mellitus. Therefore, we propose the term, "Type 3 Diabetes" to reflect this newly identified pathogenic mechanism of neurodegeneration.

CNS neurotrophins are biologically active and expressed by multiple cell types

Neutrotrophins are increasingly appreciated as potential modulators of neuronal function in the adult central nervous system (CNS). To describe the neurotrophin environment within the adult CNS, mRNA and protein expression patterns of neurotrophins-3 and -4 and of brain-derived neurotrophin were investigated in adult rat spinal cord and brain. Co-localization studies with CNS cell type-specific markers demonstrates that multiple cell types, including both neurons and glia, express these neurotrophins in the normal adult CNS. Although widely implicated in important CNS functions such as synaptic plasticity, biological activity of endogenous CNS neurotrophins has not been directly demonstrated. With a sensitive neurite outgrowth bioassay we demonstrate that CNS neurotrophins elicit neurite outgrowth and are biologically active. Moreover, antibody-blocking studies suggest that these three neurotrophins may comprise the bulk of adult CNS neurotrophic activity.

Alpha-Synuclein Expression in the Developing Human Brain

Alpha (alpha)-synuclein is a presynaptic protein, abnormal expression of which has been associated with neurodegenerative and neoplastic diseases. It is abundant in the developing vertebrate central nervous system (CNS), but less is known about its developmental expression in the human CNS. Immunohistochemical expression of alpha-synuclein was studied in 39 fetal, perinatal, pediatric, and adolescent brains. Perikaryal expression of alpha-synuclein is observed as early as 11-wk gestation in the cortical plate. Several discrete neuronal groups in the hippocampus, basal ganglia, and brain stem express perikaryal alpha-synuclein by 20-wk gestation, persisting through the first few years of life. In the cerebellum, alpha-synuclein is present by 21-wk gestation and persists into adult life as a coarse granular neuropil reaction product in the internal granular layer, and as a diffuse neuropil "blush" in the molecular layer. The germinal matrix, glia, endothelial cells, external granular layer, Pukinje cells, and dentate neurons are consistently negative for alpha-synuclein. We conclude that alpha-synuclein is expressed very early in human gestation, and that its distribution and temporal sequence of expression varies in discrete neuronal groups. Perikaryal alpha-synuclein starts disappearing from the neuronal cytosol in early childhood, and only the neuropil retains immunoreactivity into adulthood. The reappearance of alpha-synuclein in the adult neuronal cytosol in certain disease processes may represent reemergence of cues from an earlier developmental stage as part of a stress response.

The angiotensin I converting enzyme insertion/deletion polymorphism influences therapeutic outcome in major depressed women, but not in men

Angiotensin converting enzyme (ACE) is expressed in the central nervous system (CNS), where its primary function comprises degradation of neuropeptides including substance P (SP). Because of the possible antidepressant effects of SP antagonists, the influence of SP on both pathophysiology and mitigation of depression has been hypothesized. It was shown that ACE plasma concentration is determined by an insertion/deletion (I/D) polymorphism represented by the presence or absence of a 287 bp DNA fragment within the ACE gene. Because the D allele was associated with higher ACE levels this may have a positive impact on the therapeutic efficacy of antidepressant treatment. Thus, variations in CNS expression of ACE might influence the response to various antidepressant therapies. We could show a divergent clinical outcome in relation to different genotypes in 313 depressed patients who were treated with various antidepressants. A lower HAM-D17 score after 4 weeks of treatment in D/D and I/D in comparison to I/I genotypes was detected; the duration of hospitalization was shorter in D allele carriers. The D allele seems to be a predictor for a faster onset of different antidepressant therapies. The patients' gender influences these outcome effects significantly. After subdivision of the patients according to their gender only female patients contributed significantly to the genotype dependent therapeutic outcome. Our investigation gives the first hint that the speed of onset of antidepressant therapies may be dependent on both variants of the ACE genes and the gender of the patients.

The angiotensin I converting enzyme insertion/deletion polymorphism influences therapeutic outcome in major depressed women, but not in men

Angiotensin converting enzyme (ACE) is expressed in the central nervous system (CNS), where its primary function comprises degradation of neuropeptides including substance P (SP). Because of the possible antidepressant effects of SP antagonists, the influence of SP on both pathophysiology and mitigation of depression has been hypothesized. It was shown that ACE plasma concentration is determined by an insertion/deletion (I/D) polymorphism represented by the presence or absence of a 287 bp DNA fragment within the ACE gene. Because the D allele was associated with higher ACE levels this may have a positive impact on the therapeutic efficacy of antidepressant treatment. Thus, variations in CNS expression of ACE might influence the response to various antidepressant therapies. We could show a divergent clinical outcome in relation to different genotypes in 313 depressed patients who were treated with various antidepressants. A lower HAM-D17 score after 4 weeks of treatment in D/D and I/D in comparison to I/I genotypes was detected; the duration of hospitalization was shorter in D allele carriers. The D allele seems to be a predictor for a faster onset of different antidepressant therapies. The patients' gender influences these outcome effects significantly. After subdivision of the patients according to their gender only female patients contributed significantly to the genotype dependent therapeutic outcome. Our investigation gives the first hint that the speed of onset of antidepressant therapies may be dependent on both variants of the ACE genes and the gender of the patients.

Conserved cis-regulatory modules mediate complex neural expression patterns of the eyeless gene in the Drosophila brain

The Drosophila Pax-6 homologs eyeless ( ey) and twin of eyeless ( toy) are expressed in the eyes and in the central nervous system (CNS). In addition to the pivotal functions in eye development, previous studies revealed that ey also plays important roles in axonal development of the mushroom bodies, centers for associative learning and memory. It has been reported that a second intron enhancer that contains several Pax-6 binding sites mainly controls the eye-specific expression, but the DNA sequences that control CNS expression are unknown. In this work, we have dissected transcriptional enhancer elements of the ey gene that are required for the CNS expression in various developmental stages. We first show that CNS expression is independent of the eye-specific enhancer of the second intron. By systematic reporter studies, we have identified several discrete DNA elements in the 5′ upstream region and in the second intron that cooperatively interact to generate most of the ey expression pattern in the CNS. DNA sequence comparison between the ey genes of distant Drosophila species has identified conserved modules that might be bound by the upstream regulatory factors of the ey gene in CNS development. Furthermore, by RNA interference and mutant studies, we show that ey expression in the brain is independent of the activity of toy and ey itself whereas in the eye primordia it requires both, supporting the notion that ey and toy are regulated by parallel and independent regulatory cascades in brain development.

Role of the JAK/STAT signal transduction pathway in the regulation of gene expression in CNS.

Over the last 20 years the JAK/STAT signal transduction pathway has been extensively studied. An enormous amount of data on different cell signal transduction pathways is now available. The JAK/STAT signal transduction pathway is one of the intracellular signaling pathways activated by cytokines and growth factors that was first studied in the hematopoietic system, but recent data demonstrate that this signal transduction is also greatly utilized by other systems. The JAK/STAT pathway is a signaling cascade that links the activation of specific cell membrane receptors to nuclear gene expression. This review is focused on the role of JAK/STAT signal transduction pathway activation in the central nervous system (CNS).