Biomedical subjects

Frank R Sharp

Publications and source records attributed to Frank R Sharp.

At least 19 recordsLinked to original sources

PubMed2006

Relative expression of D3 dopamine receptor and alternative splice variant D3nf mRNA in high and low responders to novelty.

Studies in rodents suggest an important role for the D3 dopamine receptor in regulating locomotor responses to spatial novelty and psychostimulants. The D3 receptor alternatively spliced variant D3nf produces a non-dopamine binding protein that may alter D3 receptor localization by dimerizing with the full-length receptor. In the high responder/low responder (HR/LR) model, the locomotor response to an inescapable, novel spatial environment predicts individual differences in the locomotor and rewarding effects of psychostimulants. We hypothesized that individual differences in D3 receptor expression could contribute to individual differences in the locomotor response to novelty in the HR/LR model. To test this hypothesis, we screened rats for response to a novel spatial environment and analyzed brain tissue for mRNA levels of the D3 receptor and D3nf by real-time RT-PCR. The ratios of D3/D3nf mRNA in prefrontal cortex and substantia nigra/ventral tegmentum were significantly lower in HRs than in LRs. There were no differences in relative expression of D3/D3nf between HRs and LRs in nucleus accumbens. These data further support a role for the D3 dopamine receptor in behavioral responses to novelty and, given the established relationship between novelty and psychostimulant responses, suggest that the D3 receptor may be an important target for assessment of drug abuse vulnerability. Additionally, these findings are consistent with the hypothesis that alternative splicing may contribute to regulation of D3 dopamine receptor function.

Alternative Splicing↗

PubMed2006

Genomic profiles for human peripheral blood T cells, B cells, natural killer cells, monocytes, and polymorphonuclear cells: comparisons to ischemic stroke, migraine, and Tourette syndrome.

Blood genomic profiling has been applied to disorders of the blood and various organ systems including brain to elucidate disease mechanisms and identify surrogate disease markers. Since most studies have not examined specific cell types, we performed a preliminary genomic survey of major blood cell types from normal individuals using microarrays. CD4+ T cells, CD8+ T cells, CD19+ B cells, CD56+ natural killer cells, and CD14+ monocytes were negatively selected using the RosetteSep antibody cocktail, while polymorphonuclear leukocytes were separated with density gradient media. Genes differentially expressed by each cell type were identified. To demonstrate the potential use of such cell subtype-specific genomic expression data, a number of the major genes previously reported to be regulated in ischemic stroke, migraine, and Tourette syndrome are shown to be associated with distinct cell populations in blood. These specific gene expression, cell-type-related profiles will need to be confirmed in larger data sets and could be used to study these and many other neurological diseases.

B-Lymphocytes↗

PubMed2006

Bilirubin oxidation products (BOXes) and their role in cerebral vasospasm after subarachnoid hemorrhage.

Many factors have been postulated to cause delayed subarachnoid hemorrhage (SAH)-induced vasospasm, including hemoglobin, nitric oxide, endothelin, and free radicals. We propose that free radicals (because of the high levels that are produced in the blood clots surrounding blood vessels after SAH) act on bilirubin, biliverdin, and possibly heme to produce BOXes (Bilirubin OXidized Products). Bilirubin oxidation products act on vascular smooth muscle cells to produce chronic vasoconstriction and vasospasm combined with a vasculopathy because of smooth muscle cell injury. This review summarizes recent evidence that BOXes play a role in SAH-induced vasospasm. The data supporting a role for BOXes includes (1) identification of molecules in cerebrospinal fluid (CSF) of patients with vasospasm after SAH that have structures consistent with BOXes; (2) BOXes are vasoactive in vitro and mimic the biochemical actions of CSF of patients with vasospasm; (3) BOXes are vasoactive in vivo, constricting rat cerebral vessels; and (4) there is a correlation between clinical occurrence of vasospasm and BOXes concentration in our preliminary study of patients with SAH. Since oxidation of bilirubin, biliverdin, and perhaps heme is proposed to produce BOXes that contribute to vasospasm, either blocking bilirubin formation, inactivating bilirubin or BOXes, or removing all of the blood clot before vasospasm are potential treatment targets.

Animals↗

PubMed2006

Gene expression in blood changes rapidly in neutrophils and monocytes after ischemic stroke in humans: a microarray study.

Ischemic brain and peripheral white blood cells release cytokines, chemokines and other molecules that activate the peripheral white blood cells after stroke. To assess gene expression in these peripheral white blood cells, whole blood was examined using oligonucleotide microarrays in 15 patients at 2.4+/-0.5, 5 and 24 h after onset of ischemic stroke and compared with control blood samples. The 2.4-h blood samples were drawn before patients were treated either with tissue-type plasminogen activator (tPA) alone or with tPA plus Eptifibatide (the Combination approach to Lysis utilizing Eptifibatide And Recombinant tPA trial). Most genes induced in whole blood at 2 to 3 h were also induced at 5 and 24 h. Separate studies showed that the genes induced at 2 to 24 h after stroke were expressed mainly by polymorphonuclear leukocytes and to a lesser degree by monocytes. These genes included: matrix metalloproteinase 9; S100 calcium-binding proteins P, A12 and A9; coagulation factor V; arginase I; carbonic anhydrase IV; lymphocyte antigen 96 (cluster of differentiation (CD)96); monocarboxylic acid transporter (6); ets-2 (erythroblastosis virus E26 oncogene homolog 2); homeobox gene Hox 1.11; cytoskeleton-associated protein 4; N-formylpeptide receptor; ribonuclease-2; N-acetylneuraminate pyruvate lyase; BCL6; glycogen phosphorylase. The fold change of these genes varied from 1.6 to 6.8 and these 18 genes correctly classified 10/15 patients at 2.4 h, 13/15 patients at 5 h and 15/15 patients at 24 h after stroke. These data provide insights into the inflammatory responses after stroke in humans, and should be helpful in diagnosis, understanding etiology and pathogenesis, and guiding acute treatment and development of new treatments for stroke.

Adult↗

PubMed2006

The future of genomic profiling of neurological diseases using blood.

Sequencing of the human genome and new microarray technology make it possible to assess all genes on a single chip or array. Recent studies show different patterns of gene expression related to different tissues and diseases, and these patterns of gene expression are beginning to be used for diagnosis and treatment decisions in various types of lymphoid and solid malignancies. Because of obvious problems obtaining brain tissue, progress in genomics of neurological diseases has been slow. To address this, we demonstrated that different types of acute injury in rodent brain produced different patterns of gene expression in peripheral blood. These animal studies have now been extended to human studies. Two groups have shown that there are specific genomic profiles in the blood of patients after ischemic stroke that are highly sensitive and specific for predicting stroke. Other recent studies demonstrate specific genomic profiles in the blood of patients with Down syndrome, neurofibromatosis, tuberous sclerosis, Huntington disease, multiple sclerosis, Tourette syndrome, and others. In addition, data demonstrate specific profiles of gene expression in the blood related to different drugs, toxins, and infections. Although all of these studies are still preliminary basic scientific endeavors, they suggest that this approach will have clinical applications to neurological diseases in humans.

Age Factors↗

PubMed2006

Translational stroke research in the developing brain.

Preclinical animal models can help guide the development of clinical pediatric and newborn stroke trials. Data obtained using currently available models of hypoxia-ischemia and focal stroke have demonstrated the need for age-appropriate models. There are age-related differences in susceptibility of the immature brain to oxidative stress and inflammation, as well as in the rate and degree of apoptotic neuronal death. These issues need to be carefully addressed in designing future clinical trials.

Adult↗

PubMed2006

Brain genomics of intracerebral hemorrhage.

After intracerebral hemorrhage (ICH), many changes of gene transcription occur that may be important because they will contribute to understanding mechanisms of injury and recovery. Therefore, gene expression was assessed using Affymetrix microarrays in the striatum and the overlying cortex at 24 h after intracranial infusions of blood into the striatum of adult rats. Intracerebral hemorrhage regulated 369 of 8,740 transcripts as compared with saline-injected controls, with 104 regulated genes shared by the striatum and cortex. There were 108 upregulated and 126 downregulated genes in striatum, and 170 upregulated and 69 downregulated genes in the cortex. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) confirmed upregulation of IL-1-beta, Lipcortin 1 (annexin) and metallothionein 1,2, and downregulation of potassium voltage-gated channel, shaker-related subfamily, beta member 2 (Kcnab2). Of the functional groups of genes modulated by ICH, many metabolism and signal-transduction-related genes decreased in striatum but increased in adjacent cortex. In contrast, most enzyme, cytokine, chemokine, and immune response genes were upregulated in both striatum and in the cortex after ICH, likely in response to foreign proteins from the blood. A number of these genes may contribute to brain edema and cellular apoptosis caused by ICH. In addition, downregulation of growth factor pathways and the phosphatidylinositol 3-kinase (PI3K)/Akt pathway could also contribute to perihematoma cell death/apoptosis. Intracerebral hemorrhage-related downregulation of GABA-related genes and potassium channels might contribute to perihematoma cellular excitability and increased risk of post-ICH seizures. These genomic responses to ICH potentially provide new therapeutic targets for treatment.

Animals↗

PubMed2006

Genomics of brain and blood: progress and pitfalls.

Gene expression profiles in brain and blood of animals and humans can be useful for diagnosis, prognosis, and treatment of epilepsy. This article reviews recent progress and prospects for the future.

Animals↗

PubMed2006

Cerebral ischemia-hypoxia induces intravascular coagulation and autophagy.

Hypoxia is a critical factor for cell death or survival in ischemic stroke, but the pathological consequences of combined ischemia-hypoxia are not fully understood. Here we examine this issue using a modified Levine/Vannucci procedure in adult mice that consists of unilateral common carotid artery occlusion and hypoxia with tightly regulated body temperature. At the cellular level, ischemia-hypoxia produced proinflammatory cytokines and simultaneously activated both prosurvival (eg, synthesis of heat shock 70 protein, phosphorylation of ERK and AKT) and proapoptosis signaling pathways (eg, release of cytochrome c and AIF from mitochondria, cleavage of caspase-9 and -8). However, caspase-3 was not activated, and very few cells completed the apoptosis process. Instead, many damaged neurons showed features of autophagic/lysosomal cell death. At the tissue level, ischemia-hypoxia caused persistent cerebral perfusion deficits even after release of the carotid artery occlusion. These changes were associated with both platelet deposition and fibrin accumulation within the cerebral circulation and would be expected to contribute to infarction. Complementary studies in fibrinogen-deficient mice revealed that the absence of fibrin and/or secondary fibrin-mediated inflammatory processes significantly attenuated brain damage. Together, these results suggest that ischemia-hypoxia is a powerful stimulus for spontaneous coagulation leading to reperfusion deficits and autophagic/lysosomal cell death in brain.

Animals↗

PubMed2005

Atypical antipsychotics and a Src kinase inhibitor (PP1) prevent cortical injury produced by the psychomimetic, noncompetitive NMDA receptor antagonist MK-801.

Noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonists such as phencyclidine, ketamine, and MK-801 produce schizophrenia-like psychosis in humans. The same NMDA antagonists injure retrosplenial cortical neurons in adult rats. We examined the effects of atypical antipsychotics and an inhibitor of nonreceptor tyrosine kinase pp60 (Src) on the cortical injury produced by MK-801. An atypical antipsychotic (either clozapine, ziprasidone, olanzapine, quetiapine, or risperidone) or vehicle was administered to adult female Sprague-Dawley rats. PP1 (Src inhibitor), PP3 (nonfunctional analog of PP1) or vehicle (DMSO) was administered to another group of animals. After pretreatment, animals were injected with MK-801, killed 24 h after the MK-801, and injury to retrosplenial cortex assessed by neuronal Hsp70 protein expression. All atypical antipsychotics examined significantly attenuated MK-801-induced cortical damage. PP1 protected compared to vehicle, whereas PP3 did not protect. The ED50s (decrease injury by 50%) were as follows: PP1 <0.1 mg/kg; olanzapine 0.8 mg/kg; risperdal 1 mg/kg; clozapine 3 mg/kg; ziprasidone 32 mg/kg; and quetiapine 45 mg/kg. The data show that the atypical antipsychotics tested as well as a Src kinase inhibitor prevent the injury produced by the psychomimetic MK-801, and the potency of the atypical antipsychotics for preventing cortical injury was roughly similar to the potency of these drugs for treating psychosis in patients.

Analysis of Variance↗

PubMed2005

Blood gene expression profiling of neurologic diseases: a pilot microarray study.

BACKGROUND: Tissue gene expression profiling with arrays measures the transcription of thousands of genes. However, this approach cannot be readily used to guide clinical neurologic practice. OBJECTIVES: To determine whether clinical neurologic diseases are associated with unique patterns of up- and down-regulated genes in whole blood and to explore the possibility of using peripheral blood as a surrogate tissue in these diseases. DESIGN: Case-control study. SETTING: University-based pediatric and adult neurology clinics. PARTICIPANTS: Patients with neurofibromatosis type 1, epilepsy, or Tourette syndrome diagnosed using traditional clinical criteria; controls without disease; and controls with neurologic disease. MAIN OUTCOME MEASURE: Blood gene expression levels of greater than 12,000 genes, measured using U95A arrays. RESULTS: Neurofibromatosis type 1 and childhood epilepsy treated with carbamazepine or valproic acid are associated with distinct patterns of blood gene expression. Patients with valproic acid-responsive vs valproic acid-refractory epilepsy formed distinct subclusters. Tourette syndrome was characterized by several gene expression clusters. In 1 cluster, 6 genes-all associated with immune cell function-were overexpressed. CONCLUSION: Blood gene expression profiling can provide surrogate markers for neurologic diseases without obvious blood phenotypes.

Adolescent↗

PubMed2005

Heat shock proteins and neuromuscular disease.

The heat shock proteins are families of proteins with known activities that include chaperoning nascent peptides within the cell and cytoprotection. Most work on the nervous system has related to the role of heat shock proteins in neuroprotection from either hypoxic-ischemic or traumatic injury. The role of these proteins during normal physiological activity and injury is still under investigation. Heat shock proteins in neuromuscular disease have been investigated to some extent but were largely neglected until recently. The goal of this review is to summarize the evidence linking heat shock proteins with neuromuscular disease and to provide some insight into the roles or functions of these proteins in disease states.

Animals↗

PubMed2005

Repeated cocaine administration induces gene expression changes through the dopamine D1 receptors.

Drug addiction involves compulsive drug-seeking and drug-taking despite known adverse consequences. The enduring nature of drug addiction suggests that repeated exposure to abused drugs leads to stable alterations that likely involve changes in gene expression in the brain. The dopamine D1 receptor has been shown to mediate the long-term behavioral effects of cocaine. To examine how the persistent behavioral effects of cocaine correlate with underlying changes in gene expression, we have used D1 receptor mutant and wild-type mice to identify chronic cocaine-induced gene expression changes mediated via the D1 receptors. We focused on the caudoputamen and nucleus accumbens, two key brain regions that mediate the long-term effects of cocaine. Our analyses demonstrate that repeated cocaine administration induces changes in the expression of 109 genes, including those encoding the stromal cell-derived factor I, insulin-like growth factor binding protein 6, sigma 1 receptor, regulators of G-protein signaling protein 4, Wnt1 responsive Cdc42 homolog, Ca2+/calmodulin-dependent protein kinase II alpha subunit, and cyclin D2, via the D1 receptors. Moreover, the seven genes contain AP-1 binding sites in their promoter regions. These results suggest that genes encoding certain extracellular factors, membrane receptors and modulators, and intracellular signaling molecules, among others, are regulated by cocaine via the D1 receptor, and these AP-1 transcription complex-regulated genes might contribute to persistent cocaine-induced behavioral changes.

Animals↗

PubMed2004

Human blood genomics: distinct profiles for gender, age and neurofibromatosis type 1.

Application of gene expression profiling to human diseases will be limited by availability of tissue samples. It was postulated that germline genetic defects affect blood cells to produce unique expression patterns. This hypothesis was addressed by using a test neurological disease-neurofibromatosis type 1 (NF1), an autosomal dominant genetic disease caused by mutations of the NF1 gene at chromosome 17q11.2. Oligonucleotide arrays were used to survey the blood gene expression pattern of 12 NF1 patients compared to 96 controls. A group of genes related to tissue remodeling, bone development and tumor suppression were down-regulated in NF1 blood samples. In addition, there were blood genomic patterns for gender and age: Y chromosome genes showing higher expression in males, indicating a gene-dosage effect; and genes related to lymphocyte functions showing higher expression in children. The results suggest that genetic mutations can be manifested at the transcriptional level in peripheral blood cells and blood gene expression profiling may be useful for studying phenotypic differences of human genetic diseases and possibly providing diagnostic and prognostic markers.

Adolescent↗

PubMed2004

Hypoxia-ischemia induces DNA synthesis without cell proliferation in dying neurons in adult rodent brain.

Recent studies suggest that postmitotic neurons can reenter the cell cycle as a prelude to apoptosis after brain injury. However, most dying neurons do not pass the G1/S-phase checkpoint to resume DNA synthesis. The specific factors that trigger abortive DNA synthesis are not characterized. Here we show that the combination of hypoxia and ischemia induces adult rodent neurons to resume DNA synthesis as indicated by incorporation of bromodeoxyuridine (BrdU) and expression of G1/S-phase cell cycle transition markers. After hypoxia-ischemia, the majority of BrdU- and neuronal nuclei (NeuN)-immunoreactive cells are also terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling (TUNEL)-stained, suggesting that they undergo apoptosis. BrdU+ neurons, labeled shortly after hypoxia-ischemia, persist for >5 d but eventually disappear by 28 d. Before disappearing, these BrdU+/NeuN+/TUNEL+ neurons express the proliferating cell marker Ki67, lose the G1-phase cyclin-dependent kinase (CDK) inhibitors p16INK4 and p27Kip1 and show induction of the late G1/S-phase CDK2 activity and phosphorylation of the retinoblastoma protein. This contrasts to kainic acid excitotoxicity and traumatic brain injury, which produce TUNEL-positive neurons without evidence of DNA synthesis or G1/S-phase cell cycle transition. These findings suggest that hypoxia-ischemia triggers neurons to reenter the cell cycle and resume apoptosis-associated DNA synthesis in brain. Our data also suggest that the demonstration of neurogenesis after brain injury requires not only BrdU uptake and mature neuronal markers but also evidence showing absence of apoptotic markers. Manipulating the aberrant apoptosis-associated DNA synthesis that occurs with hypoxia-ischemia and perhaps neurodegenerative diseases could promote neuronal survival and neurogenesis.

Adrenalectomy↗

PubMed2004

Molecular identification of the ischemic penumbra.

Review of results of experimental and clinical studies indicates that the penumbra of physiologically impaired but potentially salvageable tissue surrounding the central core of focal cerebral ischemia that develops shortly after onset of major conducting vessel occlusion is complex and dynamic with severity and duration thresholds for hypoxic stress and injury that are specific to tissue site, cell type, molecular pathway or gene expression investigated and efficiency of collateral or residual flow and reperfusion. Imaging methods that have been utilized in vivo to identify penumbra and predict response to reperfusion and other protective therapies include magnetic resonance spectroscopy, diffusion- and perfusion-MRI as well as positron emission tomography. However, resolution of focal lesions characterized by lactic acidosis or cellular edema does not predict tissue survival, and imaging thresholds for resuscitation after reperfusion have not been determined experimentally. HSP-70 stress protein induction represents an endogenous protective mechanism that occurs in penumbra but not core neurones. A robust protective effect has been demonstrated during focal ischemia in transgenic mice overexpressing HSP-70 perhaps by suppressing early cytochrome c release. Delayed manganese mediated striatal neurodegeneration can be detected with T1 MRI after brief episodes of transient focal ischemia. Future studies may define endogenous cytotoxic and cytoprotective molecular penumbras that can be exploited to improve outcome after temporary focal ischemia.

Animals↗

PubMed2004

Glutamate receptor blockade attenuates glucose hypermetabolism in perihematomal brain after experimental intracerebral hemorrhage in rat.

BACKGROUND AND PURPOSE: Intracerebral hemorrhage has no effective treatment. The delayed appearance of edema, apoptosis, and inflammation in perihematomal brain suggests that these events may be targets for therapeutic intervention. To develop successful treatments, we must learn more about the effects of hemorrhage on brain tissue. In this study, we investigated the acute metabolic effects of intrastriatal hemorrhage in rat brain. METHODS: Lysed blood or saline (50 microL each) was injected into the striatum of male Sprague-Dawley rats. The rats recovered for 1 to 72 hours before injection of [14C]-2-deoxyglucose (intraperitoneally) 30 minutes before decapitation. Animals were pretreated with the N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor antagonists dizolcilpine maleate (MK-801; 1 mg/kg) or 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]quinoxaline (NBQX; 30 mg/kg), or saline vehicle. Additional animals received intrastriatal injections of glutamate (1.0 mmol/L), NMDA (1.0 mmol/L), or AMPA (0.1 mmol/L) in the place of blood. Semiquantitative autoradiographs from the brains were analyzed to determine the effects of hemorrhage on relative glucose metabolism. RESULTS: We found an acute phase of increased [14C]-2-deoxyglucose uptake in the perihematomal region that peaks 3 hours after lysed blood injection. Saline injections had no effect on striatal glucose utilization. The increased [14C]-2-deoxyglucose uptake produced by the hemorrhages was blocked by pretreatment with MK-801 and NBQX. Glutamate injections alone had no effect on striatal metabolism, whereas NMDA and AMPA injections increased [14C]-2-deoxyglucose uptake. CONCLUSIONS: The data imply that glutamate activation of NMDA or AMPA receptors increases glucose metabolism in perihematomal brain at early times after intracerebral hemorrhage. This may provide a possible target for the treatment of intracerebral hemorrhage.

Animals↗

PubMed2004

Expression of proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1beta in the brain during experimental group B streptococcal meningitis.

We performed mRNA in situ hybridization for TNF-alpha and IL-1beta from infant rats with group B streptococcal meningitis. Induction of both cytokines was seen in the ependyma and the meninges at 4 h. Both cytokines were expressed in the brain parenchyma at 12 h. Induction of IL-1beta mRNA was seen in vessels within the brain cortex. Neutrophilic infiltrate at all time points examined was minimal and could not account for the observed cytokine expression.

Animals↗