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At least 127 records · Page 7Linked to original sources
PubMed1999

MBP-1 physically associates with histone deacetylase for transcriptional repression.

MBP-1, a c-myc promoter binding protein, is a mammalian transcription factor with intriguing properties including transcriptional repression of cellular genes. Recently, we have identified and characterized two different repressor domains of MBP-1. In this report, we have demonstrated that MBP-1 physically associates with histone deacetylase (HDAC), thus promoting formation of neucleosomes that inhibit transcription. Trichostatin A, an inhibitor of histone deacetylase, significantly reduces MBP-1-mediated transcriptional repression. However, MBP-1-mediated repression on c-myc promoter is resistant to histone deacetylase activity. Our results suggest that MBP-1 represses transcription by recruiting histone deacetylase as one of the mechanisms, whereas the other mechanism is resistant to HDAC activity and probably related to direct binding of promoter sequences or interaction through yet unidentified factor.

3T3 Cells↗
PubMed1996

Elevated potassium enhances glutamate vulnerability of dopaminergic neurons developing in mesencephalic cell cultures.

This study examines the effects of high K+ concentration on the growth and development of mesencephalic cells and their glutamate vulnerability. Mesencephalic cell cultures obtained from Wistar rat embryos on the 14th gestational day were maintained for 14 days in medium with either normal (4.2 mM) or elevated (24.2 mM) potassium concentration. There was no significant difference due to various K+ concentration in cell growth and survival up to day in vitro (DIV) 13-14. In order to test the glutamate (Glu) vulnerability, cultures were treated with 100 mu M Glu for 15 min in salt solution on the DIV 3,6,8 and 13. Glu-induced neuronal damage was estimated 24 h later by measuring the neuron-specific enolase (NSE) content in the culture medium and by counting the number of tyrosine hydroxylase-immunoreactive (TH-IR) neurons. Glu had no damaging effect on the cells on DIV 3, but became pronounced beyond DIV 6. Elevated potassium concentration 24.2 mM) in the culture medium during development significantly increased neuronal vulnerability to Glu treatment, indicated by a higher increase of NSE content in the medium and by a more pronounced Glu-induced decrease of the number of TH-IR cells. The Glu-induced decrease of the number of TH-IR cells and of NSE-IR cells let us conclude that dopaminergic neurons are more vulnerable to glutamate than other neurons from mesencephalic culture.

Animals↗
PubMedDate not supplied

Electroconvulsive shock and lidocaine-induced seizures in the rat activate astrocytes as measured by glial fibrillary acidic protein.

The effects of repeated electroconvulsive shock (ECS) and/or lidocaine treatment in the rat were studied by means of biochemical markers: GFAP (glial fibrillary acidic protein), NCAM (neural cell adhesion molecule), NSE (neuron specific enolase) and D3-protein. In adult rats given daily either ECS alone or in combination with lidocaine (experiment 1) we found that ECS significantly increased the concentration of the glial marker GFAP in limbic areas: hippocampus, amygdala, and piriform cortex. The maximal increase in GFAP was found in the piriform cortex (77%). In both piriform cortex and amygdala ECS also induced a significant decrease in D3-protein (a marker of mature synapses), but no change in NCAM (especially enriched in newly formed synapses). In piriform cortex the ratio between NCAM and D3-protein was significantly increased (4%) by ECS. The lidocaine treatment, which induced seizures in some of the animals, was without significant effect on the biochemical markers. However, multiple lidocaine-induced seizures (experiment 2) were found to be associated with a significant increase in GFAP in amygdala and piriform cortex. The study shows that seizures, whether electrically or pharmacologically induced, activate astrocytes in certain brain regions. This activation is especially pronounced in the piriform cortex and may be caused by a particularly marked synaptic vulnerability and remodeling in this area, as demonstrated by the increased NCAM/D3-ratio. Synaptic remodeling and activation of astrocytes may well influence brain function and could play a role in the chain of neurobiological events underlying the clinical effects of electroconvulsive therapy (ECT).

Analysis of Variance↗
PubMed1986

Tumorigenic Xenopus cells express several maternal and early embryonic mRNAs.

Recombinant cDNA libraries were constructed from poly(A)+ RNA isolated from different stages of oogenesis and embryogenesis from the clawed toad Xenopus laevis. Hybridization analyses were used to describe the accumulation of specific RNAs represented by these cDNA clones in oocytes, embryos, adult liver, a cell line derived from Xenopus borealis embryos (Xb693), and a tumorigenic substrain of that cell line (Xb693T). It was found that from 550 cDNA clones analysed, six sequences accumulate to higher titers in poly(A)+ RNA isolated from the tumorigenic cell line compared with the non-tumorigenic cell line. All six sequences were expressed at high levels during oogenesis, and the titers of three of these sequences decreased considerably during oogenesis. DNA sequencing of these three sequences followed by a computer search of protein data banks has identified them as coding for the glycolytic enzyme enolase, the ATP-ADP carrier protein, and a-tubulin.

Animals↗
PubMed1978

Altered enzymes in the free-living nematode, Turbatrix aceti, aged in the absence of fluorodeoxyuridine.

Aging of the free-living nematode, Turbatrix aceti, has been accomplished by a procedure based upon screening of the cultures every 3--4 days to remove newborn organisms. In this system, the enzymes, isocitrate lyase, phosphoglycerate kinase and enolase all show reduced specific activities with age in crude homogenates. More critically, pure enolase shows a reduced specific activity when isolated from old compared with young organisms (958 units/mg vs. 1340 units/mg, respectively). Immunotitration experiments confirm these results. Enolase from old T. aceti requires more antiserum per unit of enzyme activity than does enolase from young organisms. This relationship holds true for both crude homogenates and pure enzyme. The above results closely parallel previous results obtained using fluorodeoxyuridine to prevent reproduction of T. aceti during aging of the organisms. Therefore, use of fluorodeoxyuridine in the aging of nematodes is perfectly safe, at least with respect to results obtained with altered enzymes.

Aging↗
PubMed2006

Progestins inhibit the neuroprotective effects of estrogen in rat hippocampus.

Although estrogen has beneficial actions in brain, recent clinical trials demonstrated adverse neural effects of hormone therapy in postmenopausal women. The cause(s) of this disconnect between experimental and clinical findings may include unanticipated effects of progestins. We report that both natural progesterone and the clinical progestin medroxyprogesterone acetate block estrogen neuroprotection. These findings underscore the need to evaluate neural actions of progestins in the rational design of hormone therapy.

Analysis of Variance↗
PubMed2003

Androgens modulate neuronal vulnerability to kainate lesion.

Testosterone has been shown to have multiple beneficial effects on neuronal viability in developing and adult animals. Most often, testosterone promotes neural health indirectly via enzymatic conversion to estradiol by aromatase. Unclear is whether androgens can directly modulate vulnerability to neuronal insults in adult animals. We investigated this issue by modulating androgen status in rats prior to challenge with the excitotoxin kainate. Adult male rats were maintained in the following conditions: i) gonadectomized (GDX) to deplete endogenous androgens, ii) GDX+replacement with dihydrotestosterone (DHT) the active and non-aromatizable testosterone metabolite, iii) sham-GDX. Animals were then lesioned with kainate and surviving hippocampal neurons quantified. In the CA2/3 and hilar regions of the hippocampus, a modest lesion was observed in sham-GDX animals corresponding to approximately 25% cell loss in comparison to non-lesioned rats. The depletion of endogenous androgens by GDX significantly augmented lesion severity, consistent with the hypothesis that androgens are involved in maintaining cell viability. Importantly, DHT hormone replacement in GDX rats significantly attenuated kainate-induced neuron loss in CA2/3, suggesting direct androgen neuroprotection. These results demonstrate that androgens act as endogenous modulators of neuron viability, a function that may be compromised in aging men as a consequence of normal, age-related androgen depletion.

Androgens↗
PubMed2006

Differential expression of basic fibroblast growth factor-2 in the developing rat brain.

Basic fibroblast growth factor-2 is a trophic molecule involved in a number of functions within the CNS, including the regulation of CNS responses to injury. Prior studies suggest that rats recover differently from injury inflicted to different regions and at different ages throughout development, and that basic fibroblast growth factor-2 may, at least in part, underlie this phenomenon. In the present study, we describe the distribution of basic fibroblast growth factor-2 at postnatal days 0, 2, 6, 10, 12, 14, 18, 21 and 30 in the indusium griseum, the external capsule, the hippocampus, the medial prefrontal cortex, the motor cortex, the rostral migratory stream, and the subventricular zone. Our results suggest a differential temporal and spatial expression of basic fibroblast growth factor-2 throughout development, which may explain the differential recovery observed from cortical lesions inflicted at different time points after birth.

Animals↗
PubMed1990

Histopathologic verification of Verhoeff's 1918 irradiation cure of retinoblastoma.

The authors studied an eye obtained postmortem from the first patient with a successfully irradiated retinoblastoma. This patient, first treated by Verhoeff in 1917, had been followed for 71 years. Ophthalmoscopy disclosed a depressed chorioretinal scar, approximately 3 disc diameters (DD) in size, with baring of the sclera temporal to the macula. This case is historically significant, in that Verhoeff and Reese debated as to whether this tumor regressed spontaneously or secondary to irradiation. Results of histopathologic, immunohistochemical, and ultrastructural examination showed a chorioretinal, neuroglial scar without evidence of calcification, necrotic tumor cells, or residual retinoblastoma. The authors discuss the clinical and histopathologic findings in spontaneously regressed retinoblastoma, retinoma/retinocytoma, and irradiated retinoblastoma. They believe that Verhoeff was correct in his belief that x-ray therapy had cured this patient.

Aged↗
PubMed1996

Structural and mechanistic studies of enolase.

The high-resolution structure of yeast enolase cocrystallized with its equilibrium mixture of substrate and product reveals the stereochemistry of substrate/product binding and therefore the groups responsible for acid/base catalysis and stabilization of the enolate intermediate. Expression and characterization of site-specific mutant forms of the enzyme have confirmed the roles of amino acid side chains in the catalysis of the first and second steps of the reaction. Coordination of both required magnesium ions to the carboxylate of the substrate/product indicates a role for these cations in stabilization of the intermediate.

Binding Sites↗
PubMed1991

Inhibition of enolase: the crystal structures of enolase-Ca2(+)- 2-phosphoglycerate and enolase-Zn2(+)-phosphoglycolate complexes at 2.2-A resolution.

Enolase is a metalloenzyme which catalyzes the elimination of H2O from 2-phosphoglyceric acid (PGA) to form phosphoenolpyruvate (PEP). Mg2+ and Zn2+ are cofactors which strongly bind and activate the enzyme. Ca2+ also binds strongly but does not produce activity. Phosphoglycolate (PG) is a competitive inhibitor of enolase. The structures of two inhibitory ternary complexes: yeast enolase-Ca2(+)-PGA and yeast enolase-Zn2(+)-PG, were determined by X-ray diffraction to 2.2-A resolution and were refined by crystallographic least-squares to R = 14.8% and 15.7%, respectively, with good geometries of the models. These structures are compared with the structure of the precatalytic ternary complex enolase-Mg2(+)-PGA/PEP (Lebioda & Stec, 1991). In the complex enolase-Ca2(+)-PGA, the PGA molecule coordinates to the Ca2+ ion with the hydroxyl group, as in the precatalytic complex. The conformation of the PGA molecule is however different. In the active complex, the organic part of the PGA molecule is planar, similar to the product. In the inhibitory complex, the carboxylic group is in an orthonormal conformation. In the inhibitory complex enolase-Zn2(+)-PG, the PG molecule coordinates with the carboxylic group in a monodentate mode. In both inhibitory complexes, the conformational changes in flexible loops, which were observed in the precatalytic complex, do not take place. The lack of catalytic metal ion binding suggests that these conformational changes are necessary for the formation of the catalytic metal ion binding site.

Amino Acid Sequence↗
PubMed2006

Evolution of enzymatic activities in the enolase superfamily: N-succinylamino acid racemase and a new pathway for the irreversible conversion of D- to L-amino acids.

Members of the mechanistically diverse enolase superfamily catalyze reactions that are initiated by abstraction of the alpha-proton of a carboxylate anion to generate an enolate anion intermediate that is stabilized by coordination to a Mg2+ ion. The catalytic groups, ligands for an essential Mg2+ and acid/base catalysts, are located in the (beta/alpha)8-barrel domain of the bidomain proteins. The assigned physiological functions in the muconate lactonizing enzyme (MLE) subgroup (Lys acid/base catalysts at the ends of the second and sixth beta-strands in the barrel domain) are cycloisomerization (MLE), dehydration (o-succinylbenzoate synthase; OSBS), and epimerization (L-Ala-D/L-Glu epimerase). We previously studied a putatively promiscuous member of the MLE subgroup with uncertain physiological function from Amycolatopsis that was discovered based on its ability to catalyze the racemization of N-acylamino acids (N-acylamino acid racemase; NAAAR) but also catalyzes the OSBS reaction [OSBS/NAAAR; Palmer, D. R., Garrett, J. B., Sharma, V., Meganathan, R., Babbitt, P. C., and Gerlt, J. A. (1999) Biochemistry 38, 4252-4258]. In this manuscript, we report functional characterization of a homologue of this protein encoded by the genome of Geobacillus kaustophilus as well as two other proteins that are encoded by the same operon, a divergent member of the Gcn5-related N-acetyltransferase (GNAT) superfamily of enzymes whose members catalyze the transfer an acyl group from an acyl-CoA donor to an amine acceptor, and a member of the M20 peptidase/carboxypeptidase G2 family. We determined that the member of the GNAT superfamily is succinyl-CoA:D-amino acid N-succinyltransferase, the member of the enolase superfamily is N-succinylamino acid racemase (NSAR), and the member of the M20 peptidase/carboxypeptidase G2 family is N-succinyl-L-amino acid hydrolase. We conclude that (1) these enzymes constitute a novel, irreversible pathway for the conversion of D- to L-amino acids and (2) the NSAR reaction is a new physiological function in the MLE subgroup. The NSAR is also functionally promiscuous and catalyzes an efficient OSBS reaction; intriguingly, the operon for menaquinone biosynthesis in G. kaustophilus does not encode an OSBS, raising the possibility that the NSAR is a bifunctional enzyme rather than an accidentally promiscuous enzyme.

Acyl Coenzyme A↗
PubMed1999

Quinupristin/dalfopristin attenuates the inflammatory response and reduces the concentration of neuron-specific enolase in the cerebrospinal fluid of rabbits with experimental Streptococcus pneumoniae meningitis.

The inflammatory response following initiation of antibiotic therapy and parameters of neuronal damage were compared during intravenous treatment with quinupristin/dalfopristin (100 mg/kg as either a short or a continuous infusion) and ceftriaxone (10 mg/kg/h) in a rabbit model of Streptococcus pneumoniae meningitis. With both modes of administration, quinupristin/dalfopristin was less bactericidal than ceftriaxone. However, the concentration of proinflammatory cell wall components (lipoteichoic acid (LTA) and teichoic acid (TA)) and the activity of tumour necrosis factor (TNF) in cerebrospinal fluid (CSF) were significantly lower in the two quinupristin/dalfopristin groups than in ceftriaxone-treated rabbits. The median LTA/TA concentrations (25th/75th percentiles) were as follows: (i) 14 h after infection: 133 (72/155) ng/mL for continuous infusion of quinupristin/dalfopristin and 193 (91/308) ng/mL for short duration infusion, compared with 455 (274/2042) ng/mL for ceftriaxone (P = 0.002 and 0.02 respectively); (ii) 17 h after infection: 116 (60/368) ng/mL for continuous infusion of quinupristin/dalfopristin and 117 (41/247) ng/mL for short duration infusion, compared with 694 (156/2173) ng/mL for ceftriaxone (P = 0.04 and 0.03 respectively). Fourteen hours after infection the median TNF activity (25th/75th percentiles) was 0.2 (0.1/1.9) U/mL for continuous infusion of quinupristin/dalfopristin and 0.1 (0.01/3.5) U/mL for short duration infusion, compared with 30 (4.6/180) U/mL for ceftriaxone (P = 0.02 for each comparison); 17 h after infection the TNF activity was 2.8 (0.2/11) U/mL (continuous infusion of quinupristin/dalfopristin) and 0.1 (0.04/6.1) U/mL (short duration infusion), compared with 48.6 (18/169) U/mL for ceftriaxone (P = 0.002 and 0.001). The concentration of neuron-specific enolase (NSE) 24 h after infection was significantly lower in animals treated with quinupristin/dalfopristin: 4.6 (3.3/5.7) microg/L (continuous infusion) and 3.6 (2.9/4.7) microg/L (short duration infusion) than in those treated with ceftriaxone (17.7 (8.8/78.2) microg/L) (P = 0.03 and 0.009 respectively). In conclusion, antibiotic treatment with quinupristin/dalfopristin attenuated the inflammatory response within the subarachnoid space after initiation of antibiotic therapy. The concentration of NSE in the CSF, taken as a measure of neuronal damage, was lower in quinupristin/dalfopristin-treated rabbits than in ceftriaxone-treated rabbits.

Animals↗
PubMed1995

Functionally essential, invariant glutamate near the C-terminus of strand beta 5 in various (alpha/beta)8-barrel enzymes as a possible indicator of their evolutionary relatedness.

Twelve different (alpha/beta)8-barrel enzymes belonging to three structurally distinct families were found to contain, near the C-terminus of their strand beta 5, a conserved invariant glutamic acid residue that plays an important functional role in each of these enzymes. The search was based on the idea that a conserved sequence region of an (alpha/beta)8-barrel enzyme should be more or less conserved also in the equivalent part of the structure of the other enzymes with this folding motif owing to their mutual evolutionary relatedness. For this purpose, the sequence region around the well conserved fifth beta-strand of alpha-amylase containing catalytic glutamate (Glu230, Aspergillus oryzae alpha-amylase numbering), was used as the sequence-structural template. The isolated sequence stretches of the 12 (alpha/beta)8-barrels are discussed from both the sequence-structural and the evolutionary point of view, the invariant glutamate residue being proposed to be a joining feature of the studied group of enzymes remaining from their ancestral (alpha/beta)8-barrel.

Amino Acid Sequence↗
PubMed2004

Lesion-dependent regulation of transgene expression in the rat brain using a human glial fibrillary acidic protein-lentiviral vector.

The ability to regulate transgene expression will be crucial for development of gene therapy to the brain. The most commonly used systems are based on a transactivator in combination with a drug, e.g. the tetracycline-regulated system. Here we describe a different method of transgene regulation by the use of the human glial fibrillary acidic protein (GFAP) promoter. We constructed a lentiviral vector that directs transgene expression to astrocytes. Using toxin-induced lesions we investigated to what extent transgene expression could be regulated in accordance with the activation of the endogenous GFAP gene. In animals receiving excitotoxic lesions of the striatum we detected an eightfold increase of green fluorescent protein (GFP)-expressing cells. The vast majority of these cells did not divide, suggesting that the transgene was indeed regulated in a similar fashion as the endogenous GFAP gene. This finding will lead to the development of lentiviral vectors with autoregulatory capacities that may be very useful for gene therapy to the brain.

Animals↗
PubMed2004

Striatal neurons in striatal grafts are derived from both post-mitotic cells and dividing progenitors.

Transplants of embryonic striatal tissue are characteristically heterogeneous, containing patches (P-zones) of striatal medium spiny projection neurons. It is not yet known how this morphology develops, and whether the striatal neurons in the grafts are derived from post-mitotic neuroblasts in the embryonic brain or from striatal progenitors that continue to divide after transplantation. To address this question we labelled dividing cells in the transplants with bromodeoxyuridine (BrdU), either prior to or after transplantation into the adult lesioned rat striatum. Cells for transplantation were either pre-labelled in utero by intraperitoneal (i.p.) injections of BrdU, or post-labelled after transplantation by i.p. injections to the hosts. Either two or six months after transplantation the brains were processed using double immunohistochemical techniques to detect BrdU and calbindin-positive neurons in the transplants. In the transplants pre-labelled with BrdU, approximately 30% of calbindin-positive cells were heavily labelled with BrdU, suggesting these had undergone a final division prior to transplantation. In transplants where cells had been labelled post-transplantation, approximately 17% of calbindin cells were heavily BrdU labelled. These results suggest that whereas a proportion of striatal medium spiny neurons in the striatal grafts were post-mitotic at the time of transplantation, other striatal progenitor cells can continue to divide after transplantation, and then complete an appropriate neuronal maturation programme in the adult host brain environment.

Acetylcholinesterase↗
PubMed2005

Disruption of the neurogenic potential of the dentate gyrus in a mouse model of temporal lobe epilepsy with focal seizures.

Adult hippocampal neurogenesis is enhanced in response to multiple stimuli including seizures. However, the relationship between neurogenesis and the development of temporal lobe epilepsy (TLE) remains unclear. Unilateral intrahippocampal injection of kainate in adult mice models the morphological characteristics (e.g. neuronal loss, gliosis, granule cell dispersion and hypertrophy) and occurrence of chronic, spontaneous recurrent partial seizures observed in human TLE. We investigated the influence of a kainate-induced epileptogenic focus on hippocampal neurogenesis, comparing neural stem cell proliferation following status epilepticus and spontaneous recurrent partial seizures. Cell proliferation in the subgranular zone was transiently increased bilaterally after kainate treatment. As a result, neurogenesis was stimulated in the contralateral dentate gyrus. In contrast, the epileptic hippocampus exhibited a strongly reduced neurogenic potential, even after onset of spontaneous recurrent partial seizures, possibly due to an alteration of the neurogenic niche in the subgranular zone. These results show that neurogenesis does not contribute to the formation of the epileptic focus and may be affected when dispersion of dentate gyrus granule cells occurs. Therefore, in patients with TLE, hippocampal sclerosis and granule cell dispersion may play a significant role in disrupting the potential for hippocampal neurogenesis.

Animals↗