Gregory Konat - Academia.edu (original) (raw)

Papers by Gregory Konat

Research paper thumbnail of Nucleotides induce higher order chromatin degradation

Molecular and Cellular Biochemistry, 2006

Higher order chromatin degradation (HOCD) is a stepwise dismantling of the genome through the exc... more Higher order chromatin degradation (HOCD) is a stepwise dismantling of the genome through the excision of chromatin loops and their oligomers at matrix attachment regions (MARs) during the early stages of programmed cell death. Although HOCD ultimately leads to the inactivation of the genome and cell death, a partial HOCD in cells receiving sublethal signals may result in the loss of genetic stability leading to neoplasia, degeneration, and aging. The present study was undertaken to determine the role of protein poly(ADP-ribosyl)ation in HOCD. Nuclei isolated from rat glioma C6 cells were able to carry poly(ADP-ribosyl)ation as assessed by the incorporation of (32)P-NAD(+) into TCA-insoluble fraction. Under the same experimental conditions, millimolar NAD(+) induced rapid HOCD in nuclei. However, while poly(ADP-ribosyl)ation was totally abrogated by specific inhibitor, benzamide, NAD(+)-induced HOCD was unaffected. Benzamide also failed to inhibit HOCD induced by H(2)O(2) exposure in intact cells. These results indicate that HOCD is not mediated through chromatin poly(ADP-ribosyl)ation, and that NAD(+) activates MAR-associated endonuclease or facilitates the access of the enzyme to DNA by other mechanisms. Furthermore, other nucleotides including NADP(+), ATP, UTP, GTP, and CTP were also found to induce HOCD in isolated nuclei indicating that HOCD is controlled by nucleotide-related ligands.

Research paper thumbnail of Kinetics of inflammatory response of astrocytes induced by TLR 3 and TLR4 ligation

Journal of Neuroscience Research, 2006

Toll-like receptors (TLRs) are sentinels of the innate immune system that recognize an array of e... more Toll-like receptors (TLRs) are sentinels of the innate immune system that recognize an array of exogenous and endogenous pathogenic molecules. The ligation of the receptors triggers inflammatory response necessary for pathogen elimination and for the healing process. In the present study we examined inflammatory response of astrocytes elicited by the ligation of TLR3 and TLR4. Astrocytic cultures established from newborn rat brains were exposed to double stranded RNA (dsRNA) and lipopolysaccharide (LPS), the ligands for TLR3 and TLR4, respectively. The expression of cytokine genes was determined by RNase protection assay, and the generation of nitric oxide (NO) was measured by Griess technique. Both ligands upregulated the expression of several cytokines (i.e., IL-1a, IL-1b, IL-6, TNFa, GM-CSF, LTb, and TGFb3) and downregulated the expression of MIF, but have no effect on the expression of IL-2, IL-3, IL-4, IL-5, IL-10, TGFb1, TGFb2, TNFb, and IFNg. Although dsRNA upregulated the expression of IFNb, LPS did not indicating that the TRIF-dependent branch of TLR4 signaling is inactive in astrocytes. Proinflammatory response as seen from upregulated cytokine expression and NO generation reached a peak within the first day of exposure, and was subsequently abrogated. The cells also became refractory to subsequent stimulation by the ligands indicating the existence of negative feedback mechanisms that control proinflammatory response in astrocytes. V

Research paper thumbnail of Statins enhance toll-like receptor 4-mediated cytokine gene expression in astrocytes: Implication of Rho proteins in negative feedback regulation

Journal of Neuroscience Research, 2008

Toll-like receptors (TLRs) are sentinels of innate immunity that recognize pathogenic molecules a... more Toll-like receptors (TLRs) are sentinels of innate immunity that recognize pathogenic molecules and trigger inflammatory response. Because inflammatory mediators are detrimental to the host, the TLR response is regulated by feedback inhibition. Statins, the inhibitors of isoprenoid biosynthesis, have been shown to be potent modulators of TLR activity, and this modulation may provide insight regarding mechanisms of the feedback inhibition. In the present study, we examined feedback mechanisms that regulate TLR4 activity in astrocytes using statins to perturb postligational signaling. Astrocytic cultures established from newborn rat brains were exposed to lipopolysaccharide (LPS), the ligand for TLR4. The up-regulation of expression of genes encoding interleukin (IL)-1b, IL-6, and tumor necrosis factor-a (TNFa) was determined by real-time RT-PCR. Pretreatment of the cells with either atorvastatin or simvastatin enhanced the LPS-induced up-regulation of cytokine gene expression. The most profound enhancement of approximately 17-fold was observed for the Il-6 gene. The enhancements for the Tnfa and Il-1b genes were approximately 5-and 3.5-fold, respectively. Mevalonate fully reversed the effects of statins, indicating that these drugs act through the inhibition of isoprenoid synthesis. The inhibition of protein geranylgeranylation, but not protein farnesylation, mimicked the effects of statins, strongly indicating that the enhancement is mediated by the Rho proteins. In support of this notion, pretreatment of cells with toxin B, a specific inhibitor of the Rho proteins, also enhanced LPS-triggered up-regulation of the cytokine genes. These results indicate that the Rho proteins are involved in the activation of negative feedback inhibition of TLR4 signaling in astrocytes. V

Research paper thumbnail of Acute effects of diets rich in almonds and walnuts on endothelial function

Indian heart journal

Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce ... more Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce cardiovascular disease (CVD) risk, by changing vascular inflammation and improving endothelial dysfunction. The objective of the study was to evaluate the acute effects of two different diets, one containing walnuts and the other almonds on endothelial function. Twenty-seven overweight volunteers underwent a randomized 2-period, crossover, controlled intervention study. The subjects were given either walnut or almond diets which varied in monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) content. The walnut diet provided 23.1% energy from PUFA and the almond diet provided 7.6% energy from PUFA. Endothelial function was assessed physiologically by flow-mediated dilation (FMD) and biochemically by sVCAM (soluble vascular cell adhesion molecules). The walnut diet significantly improved FMD (p=0.004) and decreased sVCAM (p=0.009) whereas the almond diet tended to improv...

Research paper thumbnail of Peripherally restricted viral challenge elevates extracellular glutamate and enhances synaptic transmission in the hippocampus

Journal of Neurochemistry, 2016

Peripheral infections increase the propensity and severity of seizures in susceptible populations... more Peripheral infections increase the propensity and severity of seizures in susceptible populations. We have previously shown that intraperitoneal injection of a viral mimic, polyinosinicpolycytidylic acid (PIC), elicits hypersusceptibility of mice to kainic acid (KA)-induced seizures. This study was undertaken to determine whether this seizure hypersusceptibility entails alterations in glutamate signaling. Female C57BL/6 mice were intraperitoneally injected with PIC, and after 24 h, glutamate homeostasis in the hippocampus was monitored using the enzyme-based microelectrode arrays. PIC challenge robustly increased the level of resting extracellular glutamate. While presynaptic potassium-evoked glutamate release was not affected, glutamate uptake was profoundly impaired and non-vesicular glutamate release was augmented, indicating functional alterations of astrocytes. Electrophysiological examination of hippocampal slices from PIC-challenged mice revealed a several fold increase in the basal synaptic transmission as compared to control slices. PIC challenge also increased the probability of pre-synaptic glutamate release as seen from a reduction of paired-pulse facilitation and synaptic plasticity as seen from an enhancement of long-term potentiation. Altogether, our results implicate a dysregulation of astrocytic glutamate metabolism and an alteration of excitatory synaptic transmission as the underlying mechanism for the development of hippocampal hyperexcitability, and consequently seizure hypersusceptibility following peripheral PIC challenge.

Research paper thumbnail of Comprar Signaling by Toll-Like Receptors | Gregory W. Konat | 9781420043181 | Informa Healthcare

Http Www Libreriasaulamedica Com, 2008

Tienda online donde Comprar Signaling by Toll-Like Receptors al precio 150,65 € de Gregory W. Kon... more Tienda online donde Comprar Signaling by Toll-Like Receptors al precio 150,65 € de Gregory W. Konat, tienda de Libros de Medicina, Libros de Alergologia e Inmunologia - Inmunologia

Research paper thumbnail of Generation of high efficiency single-strand DNA hybridization probe by PCR

Research paper thumbnail of Acute effects of diets rich in almonds and walnuts on endothelial function

Indian heart journal

Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce ... more Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce cardiovascular disease (CVD) risk, by changing vascular inflammation and improving endothelial dysfunction. The objective of the study was to evaluate the acute effects of two different diets, one containing walnuts and the other almonds on endothelial function. Twenty-seven overweight volunteers underwent a randomized 2-period, crossover, controlled intervention study. The subjects were given either walnut or almond diets which varied in monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) content. The walnut diet provided 23.1% energy from PUFA and the almond diet provided 7.6% energy from PUFA. Endothelial function was assessed physiologically by flow-mediated dilation (FMD) and biochemically by sVCAM (soluble vascular cell adhesion molecules). The walnut diet significantly improved FMD (p=0.004) and decreased sVCAM (p=0.009) whereas the almond diet tended to improv...

Research paper thumbnail of Peripherally restricted viral challenge elevates extracellular glutamate and enhances synaptic transmission in the hippocampus

Journal of Neurochemistry, 2016

Peripheral infections increase the propensity and severity of seizures in susceptible populations... more Peripheral infections increase the propensity and severity of seizures in susceptible populations. We have previously shown that intraperitoneal injection of a viral mimic, polyinosinicpolycytidylic acid (PIC), elicits hypersusceptibility of mice to kainic acid (KA)-induced seizures. This study was undertaken to determine whether this seizure hypersusceptibility entails alterations in glutamate signaling. Female C57BL/6 mice were intraperitoneally injected with PIC, and after 24 h, glutamate homeostasis in the hippocampus was monitored using the enzyme-based microelectrode arrays. PIC challenge robustly increased the level of resting extracellular glutamate. While presynaptic potassium-evoked glutamate release was not affected, glutamate uptake was profoundly impaired and non-vesicular glutamate release was augmented, indicating functional alterations of astrocytes. Electrophysiological examination of hippocampal slices from PIC-challenged mice revealed a several fold increase in the basal synaptic transmission as compared to control slices. PIC challenge also increased the probability of pre-synaptic glutamate release as seen from a reduction of paired-pulse facilitation and synaptic plasticity as seen from an enhancement of long-term potentiation. Altogether, our results implicate a dysregulation of astrocytic glutamate metabolism and an alteration of excitatory synaptic transmission as the underlying mechanism for the development of hippocampal hyperexcitability, and consequently seizure hypersusceptibility following peripheral PIC challenge.

Research paper thumbnail of Cerebral Response to Peripheral Challenge with a Viral Mimetic

Neurochemical research, Jan 2, 2015

It has been well established that peripheral inflammation resulting from microbial infections pro... more It has been well established that peripheral inflammation resulting from microbial infections profoundly alters brain function. This review focuses on experimental systems that model cerebral effects of peripheral viral challenge. The most common models employ the induction of the acute phase response via intraperitoneal injection of a viral mimetic, polyinosinic-polycytidylic acid (PIC). The ensuing transient surge of blood-borne inflammatory mediators induces a "mirror" inflammatory response in the brain characterized by the upregulated expression of a plethora of genes encoding cytokines, chemokines and other inflammatory/stress proteins. These inflammatory mediators modify the activity of neuronal networks leading to a constellation of behavioral traits collectively categorized as the sickness behavior. Sickness behavior is an important protective response of the host that has evolved to enhance survival and limit the spread of infections within a population. However, ...

Research paper thumbnail of Triethyllead and cerebral development: an overview

Neurotoxicology, 1984

The immature brain is unduly vulnerable to the toxic effects of triethyllead (Et3Pb). Both brain ... more The immature brain is unduly vulnerable to the toxic effects of triethyllead (Et3Pb). Both brain growth and main developmental events in the tissue are appreciably restrained by this neurotoxin. Generally, the susceptibility of brain cells to Et3Pb appears to diminish with age. The major cellular alterations in the affected tissue include the destruction of cell processes, and swelling and vacuolization in the pericaryon. The effect of Et3Pb-induced poisoning is one of hypomyelination as seen from the prominent reduction in the content of cerebral myelin. Myelin-producing cells (oligodendrocytes) seem to be particularly vulnerable to Et3Pb relative to other components of the tissue. Furthermore, the toxin specifically hampers the process of myelin membrane assembly. The inhibitory effects of Et3Pb can be attributed to the interaction of this amphiphilic compound with cellular membranes and with the process of their biogenesis.

Research paper thumbnail of Chromatin Structure and Transcriptional Activity

Chromosomal Proteins and their Role in the Regulation of Gene Expression, 1975

Research paper thumbnail of Hypochlorite-induced white matter damage

Journal of Neurochemistry, 2008

The myelin proteolipid protein (PLP) is the major structural protein of CNS myelin, accounting fo... more The myelin proteolipid protein (PLP) is the major structural protein of CNS myelin, accounting for approximately half of total myelin protein. We studied synthesis and accumulation of myelin components for two months postnatally in PLP-null mice and age-matched controls. Accumulation of myelin, as assayed by levels of whole brain cerebroside and myelin basic protein, was normal in the knockout mice. The rate of cerebroside synthesis in the knockout mice was also normal. Myelin was isolated at several ages during development, using a standard subcellular fractionation protocol. The yield of Ôpurified myelinÕ isolated from a large particle (crude mitochondrial) fraction was reduced in PLP-null mice, but increased amounts of ÔmyelinÕ were obtained in the small particle (crude microsomal) fraction. This ÔmyelinÕ in the crude microsomal fraction was identified as such by flotation on 0.85 M sucrose and the myelin-characteristic 2 : 1 molar ratio of cholesterol to cerebroside. This suggests myelin from PLP-null mice is physically more fragile than normal myelin, and that during tissue dispersion, much more PLP-null myelin is fragmented into small vesicles than is the case for normal myelin. Three hours after intracranial injection of tritiated acetate into PLPnull mice, cerebroside in myelin isolated from the large particle fraction was at a similar specific radioactivity to that isolated from the small particle (crude microsomal) fraction, suggesting that the most recently deposited PLP-null myelin is not preferentially unstable. The increased fragility evident during tissue dispersion is indicative of an underlying structural abnormality in PLP-null myelin. Whether this inherent structural instability affects myelin metabolism is under investigation.

Research paper thumbnail of Cerebral Response to Peripheral Challenge with a Viral Mimetic

Neurochemical research, Jan 2, 2015

It has been well established that peripheral inflammation resulting from microbial infections pro... more It has been well established that peripheral inflammation resulting from microbial infections profoundly alters brain function. This review focuses on experimental systems that model cerebral effects of peripheral viral challenge. The most common models employ the induction of the acute phase response via intraperitoneal injection of a viral mimetic, polyinosinic-polycytidylic acid (PIC). The ensuing transient surge of blood-borne inflammatory mediators induces a "mirror" inflammatory response in the brain characterized by the upregulated expression of a plethora of genes encoding cytokines, chemokines and other inflammatory/stress proteins. These inflammatory mediators modify the activity of neuronal networks leading to a constellation of behavioral traits collectively categorized as the sickness behavior. Sickness behavior is an important protective response of the host that has evolved to enhance survival and limit the spread of infections within a population. However, ...

Research paper thumbnail of Randomized Trial of Neuroprotective Effects of Erythropoietin in Patients Receiving Adjuvant Chemotherapy for Breast Cancer: Positron Emission Tomography and Neuropsychological Study

PsycEXTRA Dataset

Public reporting burden for this collection of information is estimated to average 1 hour per res... more Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to any penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number.

Research paper thumbnail of Generation of high efficiency ssDNA hybridization probes by linear polymerase chain reaction (LPCR)

Scanning microscopy. Supplement, 1996

The polymerase chain reaction (PCR) methodology can be employed to produce DNA hybridization prob... more The polymerase chain reaction (PCR) methodology can be employed to produce DNA hybridization probes. The major advantages of this paradigm over other techniques include superior specific activity of the probes, the versatility of sequence selection, the ability to produce short probes, and the simplicity of the procedure. We have further improved the efficiency of PCR probes by generating single stranded (ssDNA) probes that do not reanneal with themselves in solution, and hence, their availability for the interaction with the complementary sequences of the target is profoundly increased. Protocols for 32P-dCTP labeled and digoxigenin-dUTP labeled probes have been elaborated to maximize the incorporation rate of the label as well as to provide for the production of full-length probes. The ssDNA probes may be particularly suitable for nucleic acid detection in tissues by in situ hybridization.

Research paper thumbnail of Higher Order Chromatin Degradation: Implications for Neurodegeneration

Higher order chromatin degradation (HOCD) is a hallmark of programmed cell death. HOCD is mediate... more Higher order chromatin degradation (HOCD) is a hallmark of programmed cell death. HOCD is mediated by enzymatic digestion of the DNA backbone at matrix attachment regions, and ultimately results in the excision of chromatin loops and their oligomers from chromosomes. We have recently demonstrated that hydrogen peroxide (H 2 O 2), the major mediator of oxidative stress, rapidly induces HOCD. This demonstration allowed us to characterize several kinetic features of HOCD. Moreover, H 2 O 2-induced HOCD provides a mechanistic link between oxidative stress and the pathology of neurodegeneration. Thus, in acute neurodegenerative conditions, which feature severe oxidative stress, H 2 O 2-induced HOCD efficiently dismantles the genome, and thus, irreversibly commits cells to death. In chronic neurodegenerative conditions, which feature sublethal but perennial oxidative stress, cells undergo only a partial fragmentation of the genome via H 2 O 2-induced HOCD. If unrepaired of improperly repaired, such a partial fragmentation leads to the generation and accumulation of somatic mutations that are likely to play the key role in delayed degeneration and death of neural cells.

Research paper thumbnail of A Blood Test for Multiple Sclerosis

New England Journal of Medicine, 1977

Research paper thumbnail of Peripherally restricted acute phase response to a viral mimic alters hippocampal gene expression

Metabolic Brain Disease, 2013

We have previously shown that peripherally restricted acute phase response (APR) elicited by intr... more We have previously shown that peripherally restricted acute phase response (APR) elicited by intraperitoneal (i.p.) injection of a viral mimic, polyinosinic-polycytidylic acid (PIC), renders the brain hypersusceptible to excitotoxic insult as seen from profoundly exacerbated kainic acid (KA)induced seizures. In the present study, we found that this hypersusceptibility was protracted for up to 72 h. RT-PCR profiling of hippocampal gene expression revealed rapid upregulation of 23 genes encoding cytokines, chemokines and chemokine receptors generally within 6 h after PIC challenge. The expression of most of these genes decreased by 24 h. However, two chemokine genes, i.e., Ccl19 and Cxcl13 genes, as well as two chemokine receptor genes, Ccr1 and Ccr7, remained upregulated for 72 h suggesting their possible involvement in the induction and sustenance of seizure hypersusceptibility. Also, 12 genes encoding proteins related to glutamatergic and GABAergic neurotransmission featured initial upregulation or downregulation followed by gradual normalization. The upregulation of the Gabrr3 gene remained upregulated at 72 h, congruent with its plausible role in the hypersusceptible phenotype. Moreover, the expression of ten microRNAs (miRs) was rapidly affected by PIC challenge, but their levels generally exhibited oscillating profiles over the time course of seizure hypersusceptibility. These results indicate that protracted seizure susceptibility following peripheral APR is associated with a robust polygenic response in the hippocampus.

Research paper thumbnail of Endogenous Microsomal Phospholipid Peroxidation in Mouse Brain

Journal of Neurochemistry, 1973

Release of malondialdehyde from microsomal phospholipids during incubation in vitro was studied i... more Release of malondialdehyde from microsomal phospholipids during incubation in vitro was studied in the brain cytoplasm and in the microsornes fortified by deproteinized cytosoIe. Linearity between the log of specific activity of peroxidation (malondialdehyde release) and the negative log of protein concentration was demonstrated. The boiling of microsomes significantly quenched the ability to produce malondialdehyde, while preincubation with phospholipase A completely abolished this ability. In general, aromatic and not aliphatic compounds inhibited the reaction. 1,lJ-Trichloro-2,2-bis (p-chlorophenyl) ethane (DDT) was found to be a selective inhibitor of membrane peroxidative activity. The maximal inhibition is invariable. The data obtained revealed that brain and liver microsomal phospholipid peroxidation differ in some respects and the differences are of both quantitative and qualitative nature.

Research paper thumbnail of Nucleotides induce higher order chromatin degradation

Molecular and Cellular Biochemistry, 2006

Higher order chromatin degradation (HOCD) is a stepwise dismantling of the genome through the exc... more Higher order chromatin degradation (HOCD) is a stepwise dismantling of the genome through the excision of chromatin loops and their oligomers at matrix attachment regions (MARs) during the early stages of programmed cell death. Although HOCD ultimately leads to the inactivation of the genome and cell death, a partial HOCD in cells receiving sublethal signals may result in the loss of genetic stability leading to neoplasia, degeneration, and aging. The present study was undertaken to determine the role of protein poly(ADP-ribosyl)ation in HOCD. Nuclei isolated from rat glioma C6 cells were able to carry poly(ADP-ribosyl)ation as assessed by the incorporation of (32)P-NAD(+) into TCA-insoluble fraction. Under the same experimental conditions, millimolar NAD(+) induced rapid HOCD in nuclei. However, while poly(ADP-ribosyl)ation was totally abrogated by specific inhibitor, benzamide, NAD(+)-induced HOCD was unaffected. Benzamide also failed to inhibit HOCD induced by H(2)O(2) exposure in intact cells. These results indicate that HOCD is not mediated through chromatin poly(ADP-ribosyl)ation, and that NAD(+) activates MAR-associated endonuclease or facilitates the access of the enzyme to DNA by other mechanisms. Furthermore, other nucleotides including NADP(+), ATP, UTP, GTP, and CTP were also found to induce HOCD in isolated nuclei indicating that HOCD is controlled by nucleotide-related ligands.

Research paper thumbnail of Kinetics of inflammatory response of astrocytes induced by TLR 3 and TLR4 ligation

Journal of Neuroscience Research, 2006

Toll-like receptors (TLRs) are sentinels of the innate immune system that recognize an array of e... more Toll-like receptors (TLRs) are sentinels of the innate immune system that recognize an array of exogenous and endogenous pathogenic molecules. The ligation of the receptors triggers inflammatory response necessary for pathogen elimination and for the healing process. In the present study we examined inflammatory response of astrocytes elicited by the ligation of TLR3 and TLR4. Astrocytic cultures established from newborn rat brains were exposed to double stranded RNA (dsRNA) and lipopolysaccharide (LPS), the ligands for TLR3 and TLR4, respectively. The expression of cytokine genes was determined by RNase protection assay, and the generation of nitric oxide (NO) was measured by Griess technique. Both ligands upregulated the expression of several cytokines (i.e., IL-1a, IL-1b, IL-6, TNFa, GM-CSF, LTb, and TGFb3) and downregulated the expression of MIF, but have no effect on the expression of IL-2, IL-3, IL-4, IL-5, IL-10, TGFb1, TGFb2, TNFb, and IFNg. Although dsRNA upregulated the expression of IFNb, LPS did not indicating that the TRIF-dependent branch of TLR4 signaling is inactive in astrocytes. Proinflammatory response as seen from upregulated cytokine expression and NO generation reached a peak within the first day of exposure, and was subsequently abrogated. The cells also became refractory to subsequent stimulation by the ligands indicating the existence of negative feedback mechanisms that control proinflammatory response in astrocytes. V

Research paper thumbnail of Statins enhance toll-like receptor 4-mediated cytokine gene expression in astrocytes: Implication of Rho proteins in negative feedback regulation

Journal of Neuroscience Research, 2008

Toll-like receptors (TLRs) are sentinels of innate immunity that recognize pathogenic molecules a... more Toll-like receptors (TLRs) are sentinels of innate immunity that recognize pathogenic molecules and trigger inflammatory response. Because inflammatory mediators are detrimental to the host, the TLR response is regulated by feedback inhibition. Statins, the inhibitors of isoprenoid biosynthesis, have been shown to be potent modulators of TLR activity, and this modulation may provide insight regarding mechanisms of the feedback inhibition. In the present study, we examined feedback mechanisms that regulate TLR4 activity in astrocytes using statins to perturb postligational signaling. Astrocytic cultures established from newborn rat brains were exposed to lipopolysaccharide (LPS), the ligand for TLR4. The up-regulation of expression of genes encoding interleukin (IL)-1b, IL-6, and tumor necrosis factor-a (TNFa) was determined by real-time RT-PCR. Pretreatment of the cells with either atorvastatin or simvastatin enhanced the LPS-induced up-regulation of cytokine gene expression. The most profound enhancement of approximately 17-fold was observed for the Il-6 gene. The enhancements for the Tnfa and Il-1b genes were approximately 5-and 3.5-fold, respectively. Mevalonate fully reversed the effects of statins, indicating that these drugs act through the inhibition of isoprenoid synthesis. The inhibition of protein geranylgeranylation, but not protein farnesylation, mimicked the effects of statins, strongly indicating that the enhancement is mediated by the Rho proteins. In support of this notion, pretreatment of cells with toxin B, a specific inhibitor of the Rho proteins, also enhanced LPS-triggered up-regulation of the cytokine genes. These results indicate that the Rho proteins are involved in the activation of negative feedback inhibition of TLR4 signaling in astrocytes. V

Research paper thumbnail of Acute effects of diets rich in almonds and walnuts on endothelial function

Indian heart journal

Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce ... more Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce cardiovascular disease (CVD) risk, by changing vascular inflammation and improving endothelial dysfunction. The objective of the study was to evaluate the acute effects of two different diets, one containing walnuts and the other almonds on endothelial function. Twenty-seven overweight volunteers underwent a randomized 2-period, crossover, controlled intervention study. The subjects were given either walnut or almond diets which varied in monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) content. The walnut diet provided 23.1% energy from PUFA and the almond diet provided 7.6% energy from PUFA. Endothelial function was assessed physiologically by flow-mediated dilation (FMD) and biochemically by sVCAM (soluble vascular cell adhesion molecules). The walnut diet significantly improved FMD (p=0.004) and decreased sVCAM (p=0.009) whereas the almond diet tended to improv...

Research paper thumbnail of Peripherally restricted viral challenge elevates extracellular glutamate and enhances synaptic transmission in the hippocampus

Journal of Neurochemistry, 2016

Peripheral infections increase the propensity and severity of seizures in susceptible populations... more Peripheral infections increase the propensity and severity of seizures in susceptible populations. We have previously shown that intraperitoneal injection of a viral mimic, polyinosinicpolycytidylic acid (PIC), elicits hypersusceptibility of mice to kainic acid (KA)-induced seizures. This study was undertaken to determine whether this seizure hypersusceptibility entails alterations in glutamate signaling. Female C57BL/6 mice were intraperitoneally injected with PIC, and after 24 h, glutamate homeostasis in the hippocampus was monitored using the enzyme-based microelectrode arrays. PIC challenge robustly increased the level of resting extracellular glutamate. While presynaptic potassium-evoked glutamate release was not affected, glutamate uptake was profoundly impaired and non-vesicular glutamate release was augmented, indicating functional alterations of astrocytes. Electrophysiological examination of hippocampal slices from PIC-challenged mice revealed a several fold increase in the basal synaptic transmission as compared to control slices. PIC challenge also increased the probability of pre-synaptic glutamate release as seen from a reduction of paired-pulse facilitation and synaptic plasticity as seen from an enhancement of long-term potentiation. Altogether, our results implicate a dysregulation of astrocytic glutamate metabolism and an alteration of excitatory synaptic transmission as the underlying mechanism for the development of hippocampal hyperexcitability, and consequently seizure hypersusceptibility following peripheral PIC challenge.

Research paper thumbnail of Comprar Signaling by Toll-Like Receptors | Gregory W. Konat | 9781420043181 | Informa Healthcare

Http Www Libreriasaulamedica Com, 2008

Tienda online donde Comprar Signaling by Toll-Like Receptors al precio 150,65 € de Gregory W. Kon... more Tienda online donde Comprar Signaling by Toll-Like Receptors al precio 150,65 € de Gregory W. Konat, tienda de Libros de Medicina, Libros de Alergologia e Inmunologia - Inmunologia

Research paper thumbnail of Generation of high efficiency single-strand DNA hybridization probe by PCR

Research paper thumbnail of Acute effects of diets rich in almonds and walnuts on endothelial function

Indian heart journal

Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce ... more Omega-3 fatty acids, especially alpha-linolenic acid (ALA), which are present in nuts may reduce cardiovascular disease (CVD) risk, by changing vascular inflammation and improving endothelial dysfunction. The objective of the study was to evaluate the acute effects of two different diets, one containing walnuts and the other almonds on endothelial function. Twenty-seven overweight volunteers underwent a randomized 2-period, crossover, controlled intervention study. The subjects were given either walnut or almond diets which varied in monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) content. The walnut diet provided 23.1% energy from PUFA and the almond diet provided 7.6% energy from PUFA. Endothelial function was assessed physiologically by flow-mediated dilation (FMD) and biochemically by sVCAM (soluble vascular cell adhesion molecules). The walnut diet significantly improved FMD (p=0.004) and decreased sVCAM (p=0.009) whereas the almond diet tended to improv...

Research paper thumbnail of Peripherally restricted viral challenge elevates extracellular glutamate and enhances synaptic transmission in the hippocampus

Journal of Neurochemistry, 2016

Peripheral infections increase the propensity and severity of seizures in susceptible populations... more Peripheral infections increase the propensity and severity of seizures in susceptible populations. We have previously shown that intraperitoneal injection of a viral mimic, polyinosinicpolycytidylic acid (PIC), elicits hypersusceptibility of mice to kainic acid (KA)-induced seizures. This study was undertaken to determine whether this seizure hypersusceptibility entails alterations in glutamate signaling. Female C57BL/6 mice were intraperitoneally injected with PIC, and after 24 h, glutamate homeostasis in the hippocampus was monitored using the enzyme-based microelectrode arrays. PIC challenge robustly increased the level of resting extracellular glutamate. While presynaptic potassium-evoked glutamate release was not affected, glutamate uptake was profoundly impaired and non-vesicular glutamate release was augmented, indicating functional alterations of astrocytes. Electrophysiological examination of hippocampal slices from PIC-challenged mice revealed a several fold increase in the basal synaptic transmission as compared to control slices. PIC challenge also increased the probability of pre-synaptic glutamate release as seen from a reduction of paired-pulse facilitation and synaptic plasticity as seen from an enhancement of long-term potentiation. Altogether, our results implicate a dysregulation of astrocytic glutamate metabolism and an alteration of excitatory synaptic transmission as the underlying mechanism for the development of hippocampal hyperexcitability, and consequently seizure hypersusceptibility following peripheral PIC challenge.

Research paper thumbnail of Cerebral Response to Peripheral Challenge with a Viral Mimetic

Neurochemical research, Jan 2, 2015

It has been well established that peripheral inflammation resulting from microbial infections pro... more It has been well established that peripheral inflammation resulting from microbial infections profoundly alters brain function. This review focuses on experimental systems that model cerebral effects of peripheral viral challenge. The most common models employ the induction of the acute phase response via intraperitoneal injection of a viral mimetic, polyinosinic-polycytidylic acid (PIC). The ensuing transient surge of blood-borne inflammatory mediators induces a "mirror" inflammatory response in the brain characterized by the upregulated expression of a plethora of genes encoding cytokines, chemokines and other inflammatory/stress proteins. These inflammatory mediators modify the activity of neuronal networks leading to a constellation of behavioral traits collectively categorized as the sickness behavior. Sickness behavior is an important protective response of the host that has evolved to enhance survival and limit the spread of infections within a population. However, ...

Research paper thumbnail of Triethyllead and cerebral development: an overview

Neurotoxicology, 1984

The immature brain is unduly vulnerable to the toxic effects of triethyllead (Et3Pb). Both brain ... more The immature brain is unduly vulnerable to the toxic effects of triethyllead (Et3Pb). Both brain growth and main developmental events in the tissue are appreciably restrained by this neurotoxin. Generally, the susceptibility of brain cells to Et3Pb appears to diminish with age. The major cellular alterations in the affected tissue include the destruction of cell processes, and swelling and vacuolization in the pericaryon. The effect of Et3Pb-induced poisoning is one of hypomyelination as seen from the prominent reduction in the content of cerebral myelin. Myelin-producing cells (oligodendrocytes) seem to be particularly vulnerable to Et3Pb relative to other components of the tissue. Furthermore, the toxin specifically hampers the process of myelin membrane assembly. The inhibitory effects of Et3Pb can be attributed to the interaction of this amphiphilic compound with cellular membranes and with the process of their biogenesis.

Research paper thumbnail of Chromatin Structure and Transcriptional Activity

Chromosomal Proteins and their Role in the Regulation of Gene Expression, 1975

Research paper thumbnail of Hypochlorite-induced white matter damage

Journal of Neurochemistry, 2008

The myelin proteolipid protein (PLP) is the major structural protein of CNS myelin, accounting fo... more The myelin proteolipid protein (PLP) is the major structural protein of CNS myelin, accounting for approximately half of total myelin protein. We studied synthesis and accumulation of myelin components for two months postnatally in PLP-null mice and age-matched controls. Accumulation of myelin, as assayed by levels of whole brain cerebroside and myelin basic protein, was normal in the knockout mice. The rate of cerebroside synthesis in the knockout mice was also normal. Myelin was isolated at several ages during development, using a standard subcellular fractionation protocol. The yield of Ôpurified myelinÕ isolated from a large particle (crude mitochondrial) fraction was reduced in PLP-null mice, but increased amounts of ÔmyelinÕ were obtained in the small particle (crude microsomal) fraction. This ÔmyelinÕ in the crude microsomal fraction was identified as such by flotation on 0.85 M sucrose and the myelin-characteristic 2 : 1 molar ratio of cholesterol to cerebroside. This suggests myelin from PLP-null mice is physically more fragile than normal myelin, and that during tissue dispersion, much more PLP-null myelin is fragmented into small vesicles than is the case for normal myelin. Three hours after intracranial injection of tritiated acetate into PLPnull mice, cerebroside in myelin isolated from the large particle fraction was at a similar specific radioactivity to that isolated from the small particle (crude microsomal) fraction, suggesting that the most recently deposited PLP-null myelin is not preferentially unstable. The increased fragility evident during tissue dispersion is indicative of an underlying structural abnormality in PLP-null myelin. Whether this inherent structural instability affects myelin metabolism is under investigation.

Research paper thumbnail of Cerebral Response to Peripheral Challenge with a Viral Mimetic

Neurochemical research, Jan 2, 2015

It has been well established that peripheral inflammation resulting from microbial infections pro... more It has been well established that peripheral inflammation resulting from microbial infections profoundly alters brain function. This review focuses on experimental systems that model cerebral effects of peripheral viral challenge. The most common models employ the induction of the acute phase response via intraperitoneal injection of a viral mimetic, polyinosinic-polycytidylic acid (PIC). The ensuing transient surge of blood-borne inflammatory mediators induces a "mirror" inflammatory response in the brain characterized by the upregulated expression of a plethora of genes encoding cytokines, chemokines and other inflammatory/stress proteins. These inflammatory mediators modify the activity of neuronal networks leading to a constellation of behavioral traits collectively categorized as the sickness behavior. Sickness behavior is an important protective response of the host that has evolved to enhance survival and limit the spread of infections within a population. However, ...

Research paper thumbnail of Randomized Trial of Neuroprotective Effects of Erythropoietin in Patients Receiving Adjuvant Chemotherapy for Breast Cancer: Positron Emission Tomography and Neuropsychological Study

PsycEXTRA Dataset

Public reporting burden for this collection of information is estimated to average 1 hour per res... more Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to any penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number.

Research paper thumbnail of Generation of high efficiency ssDNA hybridization probes by linear polymerase chain reaction (LPCR)

Scanning microscopy. Supplement, 1996

The polymerase chain reaction (PCR) methodology can be employed to produce DNA hybridization prob... more The polymerase chain reaction (PCR) methodology can be employed to produce DNA hybridization probes. The major advantages of this paradigm over other techniques include superior specific activity of the probes, the versatility of sequence selection, the ability to produce short probes, and the simplicity of the procedure. We have further improved the efficiency of PCR probes by generating single stranded (ssDNA) probes that do not reanneal with themselves in solution, and hence, their availability for the interaction with the complementary sequences of the target is profoundly increased. Protocols for 32P-dCTP labeled and digoxigenin-dUTP labeled probes have been elaborated to maximize the incorporation rate of the label as well as to provide for the production of full-length probes. The ssDNA probes may be particularly suitable for nucleic acid detection in tissues by in situ hybridization.

Research paper thumbnail of Higher Order Chromatin Degradation: Implications for Neurodegeneration

Higher order chromatin degradation (HOCD) is a hallmark of programmed cell death. HOCD is mediate... more Higher order chromatin degradation (HOCD) is a hallmark of programmed cell death. HOCD is mediated by enzymatic digestion of the DNA backbone at matrix attachment regions, and ultimately results in the excision of chromatin loops and their oligomers from chromosomes. We have recently demonstrated that hydrogen peroxide (H 2 O 2), the major mediator of oxidative stress, rapidly induces HOCD. This demonstration allowed us to characterize several kinetic features of HOCD. Moreover, H 2 O 2-induced HOCD provides a mechanistic link between oxidative stress and the pathology of neurodegeneration. Thus, in acute neurodegenerative conditions, which feature severe oxidative stress, H 2 O 2-induced HOCD efficiently dismantles the genome, and thus, irreversibly commits cells to death. In chronic neurodegenerative conditions, which feature sublethal but perennial oxidative stress, cells undergo only a partial fragmentation of the genome via H 2 O 2-induced HOCD. If unrepaired of improperly repaired, such a partial fragmentation leads to the generation and accumulation of somatic mutations that are likely to play the key role in delayed degeneration and death of neural cells.

Research paper thumbnail of A Blood Test for Multiple Sclerosis

New England Journal of Medicine, 1977

Research paper thumbnail of Peripherally restricted acute phase response to a viral mimic alters hippocampal gene expression

Metabolic Brain Disease, 2013

We have previously shown that peripherally restricted acute phase response (APR) elicited by intr... more We have previously shown that peripherally restricted acute phase response (APR) elicited by intraperitoneal (i.p.) injection of a viral mimic, polyinosinic-polycytidylic acid (PIC), renders the brain hypersusceptible to excitotoxic insult as seen from profoundly exacerbated kainic acid (KA)induced seizures. In the present study, we found that this hypersusceptibility was protracted for up to 72 h. RT-PCR profiling of hippocampal gene expression revealed rapid upregulation of 23 genes encoding cytokines, chemokines and chemokine receptors generally within 6 h after PIC challenge. The expression of most of these genes decreased by 24 h. However, two chemokine genes, i.e., Ccl19 and Cxcl13 genes, as well as two chemokine receptor genes, Ccr1 and Ccr7, remained upregulated for 72 h suggesting their possible involvement in the induction and sustenance of seizure hypersusceptibility. Also, 12 genes encoding proteins related to glutamatergic and GABAergic neurotransmission featured initial upregulation or downregulation followed by gradual normalization. The upregulation of the Gabrr3 gene remained upregulated at 72 h, congruent with its plausible role in the hypersusceptible phenotype. Moreover, the expression of ten microRNAs (miRs) was rapidly affected by PIC challenge, but their levels generally exhibited oscillating profiles over the time course of seizure hypersusceptibility. These results indicate that protracted seizure susceptibility following peripheral APR is associated with a robust polygenic response in the hippocampus.

Research paper thumbnail of Endogenous Microsomal Phospholipid Peroxidation in Mouse Brain

Journal of Neurochemistry, 1973

Release of malondialdehyde from microsomal phospholipids during incubation in vitro was studied i... more Release of malondialdehyde from microsomal phospholipids during incubation in vitro was studied in the brain cytoplasm and in the microsornes fortified by deproteinized cytosoIe. Linearity between the log of specific activity of peroxidation (malondialdehyde release) and the negative log of protein concentration was demonstrated. The boiling of microsomes significantly quenched the ability to produce malondialdehyde, while preincubation with phospholipase A completely abolished this ability. In general, aromatic and not aliphatic compounds inhibited the reaction. 1,lJ-Trichloro-2,2-bis (p-chlorophenyl) ethane (DDT) was found to be a selective inhibitor of membrane peroxidative activity. The maximal inhibition is invariable. The data obtained revealed that brain and liver microsomal phospholipid peroxidation differ in some respects and the differences are of both quantitative and qualitative nature.