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At least 1,099 records · Page 61Linked to original sources
PubMed2000

Regulation of brain-derived neurotrophic factor messenger RNA levels in avian hypothalamic slice cultures.

Mechanisms regulating the expression of brain-derived neurotrophic factor, a member of the neurotrophin family, have been extensively studied in the rat cerebral cortex, hippocampus and cerebellum. In contrast, little is known regarding the regulation of this growth factor in the hypothalamus. Here we present an analysis of the regulation of brain-derived neurotrophic factor messenger RNA levels in chick embryo hypothalamic slice cultures following exposure to potassium chloride, glutamate agonists and sex steroids. Following a week in chemically-defined media the tissue was depolarized by exposure to 50 mM potassium chloride for 6h, resulting in a significant 4.2-fold increase in the level of brain-derived neurotrophic factor messenger RNA. This result is consistent with studies of other brain regions. Similar 6-h acute exposures of the hypothalamic cultures to 25 microM N-methyl-D-aspartic acid, 25 microM kainic acid and 25 microM alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid also significantly increased messenger RNA levels 2.5-, 2.1- and 1.4-fold, respectively. It was previously reported that brain-derived neurotrophic factor levels within the rat cerebral cortex, olfactory bulb and hippocampus are altered by exposure to 17beta-estradiol. Here we show that in hypothalamic slice cultures neither acute nor chronic treatments with 10 and 100 nM 17beta-estradiol and 10nM testosterone significantly altered the steady-state level of this growth factor. These findings show that neuronal activity, induced by glutamate agonists and potassium chloride, can regulate brain-derived neurotrophic factor messenger RNA levels within embryonic hypothalamic slice cultures. This regulation could play a critical role in the modulation of programmed cell death and synaptic maturation during development of the hypothalamus.

Animals↗
PubMed1973

Stabilization of interferon messenger RNA activity by treatment of cells with metabolic inhibitors and lowering of the incubation temperature.

Interferon production was induced in a strain of human diploid foreskin cells with poly(I).poly(C). Cycloheximide was included in the culture medium at the time of addition of the inducer. Actinomycin D was added to the cultures 4 or 5 hr later, before the inhibition of protein synthesis was reversed at 6 hr. Thus, subsequent interferon synthesis had to be directed by messenger RNA synthesized before addition of actinomycin D. The amount of interferon produced after such treatment was about 50-fold greater than in cells induced with poly(I).poly(C) but not treated with inhibitors. Experiments using cordycepin suggested that in spite of the continued presence of the inducer the synthesis of the bulk of interferon messenger RNA was completed within the first 2 hr of exposure of cells to poly(I).poly(C) and cycloheximide. Transcription of interferon messenger RNA was apparently not affected when interferon synthesis was suppressed to various degrees by different inhibitors of protein synthesis, indicating the independence of transcription and translation. The high rate of interferon synthesis after the reversal of cycloheximide action was more sustained at 32 degrees than at 37 degrees . The rate of decrease of overall protein synthesis in cells treated with actinomycin D and then incubated either at 32 degrees or 37 degrees showed a similar dependence on incubation temperature, suggesting that the stability of messenger RNA (or of another actinomycin D-sensitive component required for protein synthesis) was greater at the lower temperature.

Carbon Radioisotopes↗
PubMed1988

Expression of ventricular-type myosin light chain messenger RNA in spontaneously hypertensive rat atria.

Using cloned DNA probes specific for two isoforms of cardiac myosin light chains (MLCs), nonphosphorylatable MLC1 and phosphorylatable, regulatory MLC2, we have observed that the MLC1 messenger RNA of ventricular type does not appear in detectable amounts in atrial cells of either normotensive Wistar-Kyoto rat strain (WKY) or spontaneously hypertensive rat strain (SHR). The messenger RNA of regulatory isoform of ventricular MLC2, on the other hand, is found in threefold excess in atria of SHR relative to that of age-matched WKY. The increased level of MLC2 messenger RNA is present even in 6-week-old SHR atria where there is no established overloading of the heart. Thus, it appears that the increased expression of the regulatory MLC2 gene in SHR atrial cells is a predetermined event, which, most likely, participates in functional adaptation of the myocardium in response to pressure overload and subsequent hypertrophy.

Animals↗
PubMed1993

Analysis of p60 and p80 tumor necrosis factor-alpha receptor messenger RNA and protein in human placentas.

Tumor necrosis factor-alpha (TNF), a pleiotrophic, multifunctional polypeptide factor, has been reported in both normal and infected human placentas. To identify potential targets for this cytokine, the cells in early and late gestation placentas and extraplacental membranes that express the two TNF receptor (TNF-R) genes, p60 and p80, were identified by using in situ hybridization and immunocytochemistry. Gestation-related, cell lineage-specific differences in steady-state levels of p60 and p80 TNF-R messenger RNA were observed. p60 TNF-R messenger RNA predominated at both early and late stages of gestation, being high in both mesenchymal and trophoblastic cell lineages. By contrast, p80 TNF-R messenger RNA was abundant only in intermittent stretches of first trimester syncytiotrophoblast and term placental mesenchymal cells. Overall, intensities of the TNF-R hybridization signals were stronger in term than in first trimester tissues. Transcription of the two TNF-R genes was confirmed by Northern blot hybridization. Translation was verified in all samples by immunohistology using polyclonal antibodies specific for the receptor proteins. p60 and p80 TNF-R proteins were identified both intracellularly and in maternal and fetal blood. Because TNF-Rs exist in both membrane-bound and soluble forms, the results of this study are consistent with the postulate that placental TNF-R have two critical functions: 1) modulation of TNF utilization by specific placental cell lineages during the course of pregnancy; and 2) protection against excessive TNF produced during infections.

Blotting, Northern↗
PubMed1999

Melanocortin-4 receptor messenger RNA expression is up-regulated in the non-damaged striatum following unilateral hypoxic-ischaemic brain injury.

Melanocortin peptides (alpha-melanocyte-stimulating hormone, adrenocorticotropin and fragments thereof) have been shown to have numerous effects on the central nervous system, including recovery from nerve injury and retention of learned behaviour, but the mechanism of action of these peptides is unknown. A family of five melanocortin receptors have recently been discovered, two of which (melanocortin-3 and melanocortin-4 receptors) have been mapped in the rat brain. We have tested the hypothesis that the expression of one or more of the messenger RNAs for three melanocortin receptors (melanocortin-3, melanocortin-4 and melanocortin-5 receptors) would be altered in rat brain following unilateral transient hypoxic-ischaemic brain injury. In this study, using in situ hybridization, we show that melanocortin-4 receptor messenger RNA was up-regulated in the striatum in the non-damaged hemisphere within 24 h after severe hypoxic-ischaemic injury compared with control brains (P<0.05). In a small group of animals, this induction was not blocked by treatment with the anticonvulsant, carbamazepine. Expression of melanocortin-3 receptor messenger RNA in the brain was not altered in this hypoxic-ischaemic injury model and melanocortin-5 receptor messenger RNA was not detected in either control or hypoxic-ischaemic injured rat brains. We hypothesize that the up-regulation of melanocortin-4 receptor messenger RNA expression in the contralateral striatum may be involved in transfer of function to the uninjured hemisphere following unilateral brain injury.

Animals↗
PubMed2004

The effects of dexamethasone and betamethasone on surfactant protein-B messenger RNA expression in human type II pneumocytes and human lung adenocarcinoma cells.

OBJECTIVE: The purpose of this study was to compare the effect of a single 48-hour exposure to betamethasone or dexamethasone in the NCI-H441 cell line and in human type II pneumocytes. STUDY DESIGN: NCI-H441 cells were exposed 48 hours to varying concentrations of betamethasone or dexamethasone (10(-10) to 10(-7) mol/L) alone or in combination with 1 mmol/L dibutyryl cyclic adenosine monophosphate. Likewise, human type II pneumocytes were exposed 48 hours to varying concentrations of betamethasone or dexamethasone (10(-9) to 10(-7) mol/L) alone or in combination with 1 mmol/L dibutyryl cyclic adenosine monophosphate. The measured outcome was the stimulatory effect on surfactant protein B gene transcription as expressed by surfactant protein B messenger RNA accumulation. The experiment was conducted 5 times in NCI-H441 cells and 6 times in type II cells, in parallel with control. Surfactant protein B messenger RNA was determined at control level and 48 hours after exposure by quantitative reverse transcription-polymerase chain reaction. RESULTS: A similar dose-dependent response in surfactant protein B messenger RNA expression was seen with both betamethasone and dexamethasone. In human type II pneumocytes, the inductive profile of surfactant protein B messenger RNA after 48-hour exposure to betamethasone or dexamethasone was similar to that seen in the NCI-H441 cells. CONCLUSION: Dexamethasone and betamethasone achieved similar dose-response patterns of surfactant protein-B expression in vitro.

Adenocarcinoma↗
PubMed1979

Analysis of the genetic complexity and abundance classes of messenger RNA in human liver and leukemic cells.

In an effort to determine the number of genes expressed as messenger RNA in disparate human tissues we have analyzed the genetic complexity of the polyribosome-associated poly(A)-containing RNA population obtained from liver and lymphoblastic leukemic cells. This was accomplished by measuring the kinetics of hybridization of mRNA to a complementary DNA probe synthesized by avian myeloblastosis virus reverse transcriptase in vitro. The results obtained from such an analysis revealed the presence of two major abundance classes of mRNA with a total genetic complexity of approximately 10,000 diverse mRNA species in both of these cell types. Diversity of mRNA species in these unrelated human cells was studied by heterologous hybridization reactions between the cDNA of one cell type and a vast excess of poly(A)-containing mRNA from another. These types of studies indicated that extensive homology (more than 80%) exists in the mRNA sequences of disparate human cell types and suggest that the vast majority of genetic information expressed as mRNA is required for the maintenance of cellular functions common to functionally different human tissues.

Avian Myeloblastosis Virus↗
PubMed1995

Hypoxemia up-regulates interleukin-8 stimulated phagocytosis of polymorphonuclear leukocytes by differential regulation of CD32w and CD35 messenger RNA expression.

BACKGROUND: The purpose of these studies was to investigate the effect of hypoxemia on interleukin-8 (IL-8) regulation of phagocytosis and surface opsonic receptor expression of whole blood polymorphonuclear leukocytes (PMNs). METHODS: Whole blood PMNs were rendered hypoxemic (PvO2 less than 15 mm Hg) or normoxic (PvO2 equals 60 to 80 mm Hg) and incubated with IL-8 (50 ng/ml) before the sequential addition of serum-opsonized fluorescent microspheres (emission wavelength, 560 nm) and fluorescein isothiocyanate-conjugated mouse anti-human CD32w or CD35 antibodies (emission wavelength, 513 nm). Concomitant two color flow cytometric analysis was then performed to measure the mean channel fluorescence (MCF) and the percentage of positive PMNs. Steady-state messenger RNA levels for CD32w and CD35 were quantitated by means of Northern blot analysis by using total RNA extracted from purified PMNs from each study group. RESULTS: Hypoxemia increased IL-8 stimulated PMN phagocytosis (percentage of positive PMNs, 38 +/- 1.5 versus 27 +/- 2.0, MCF, 5491 +/- 182 versus 4060 +/- 121 for hypoxemia plus IL-8 versus normoxia plus IL-8, respectively; p < 0.05). Hypoxemia increased IL-8 stimulated PMN expression of CD32w and CD35 (MCF CD32w, 791 +/- 105 versus 336 +/- 81; MCF CD35, 542 +/- 87 versus 254 +/- 41; p < 0.05). Under normoxic conditions IL-8 decreased messenger RNA levels for both CD32w and CD35, but under hypoxemic conditions IL-8 increased messenger RNA levels only for CD32w. CONCLUSIONS: Hypoxemia directly regulates IL-8 control of PMNs by increasing phagocytosis, receptor expression, and messenger RNA for these receptors. Studies investigating cytokine regulation of PMN function need to take into account oxygen tension when establishing the fundamental effects of cytokines on PMN physiology.

Cytokines↗
PubMed1992

Molecular biology of renin. II: Gene control by messenger RNA, transfection and transgenic studies.

PURPOSE: This review addresses the question of renin gene control by examining information from messenger RNA, transgene, transient expression and DNA-protein binding studies. CONTENTS: Although information from messenger RNA studies may be reliable for tissues such as kidney and, in mouse, submandibular gland, which have relatively high expression, more sensitive approaches, e.g. polymerase chain reaction technology, are needed for studies of renin gene regulation in other tissues. Control at the molecular level is particularly uncertain, with conflicting data suggesting that renin promoter activity depends either upon a complex interaction of positive and negative regulatory elements or upon the presence of specific trans-acting factor(s) alone, which act upon putative enhancer(s) in the renin-associated DNA. Only after renin-synthesizing cell lines are studied will a clearer picture emerge. Transgenic work has loosely defined the DNA needed as being less than several kilobases of flanking sequences, with the possibility that sequences in the gene itself, particularly in intron 1, are necessary. Such transgenic work has also shown that the renin gene can cause hypertension. CONCLUSION: Knowledge of DNA and proteins and the interactions that turn on transcription of the renin gene is still rudimentary and currently represents the major challenge in renin research.

Animals↗
PubMed1976

Messenger RNA translation in the presence of homologous and heterologous tRNA.

The effect of tRNA distribution in the cell on the rate of specific protein synthesis has been investigated. A tRNA-dependent cell-free protein-synthesizing system derived from Krebs-II ascites cells has been worked out. In this system it is possible to synthesize specific proteins (egg-white proteins or globin) by the translation of oviduct or reticulocyte messenger RNA in the presence of tRNA from homologous or heterologous tissues. The rate of translation of a give messenger RNA is optimal in the presence of tRNA from the homologous tissue. The lower level of protein synthesis obtained with an excess of heterologous tRNA can be overcome by adding the homologous tRNA. Nevertheless homologous isoaccepting species partly purified by a reversed-phase chromatography or a benzoylated DEAE-cellulose column cannot fully restore the optimal synthesis, eliminating the hypothesis according to which a particular tRNA species is involved in this phenomenon. The correct distribution of tRNA is necessary for optimal translation of a messenger RNA.

Animals↗
PubMed1993

Marked increase in cholecystokinin B receptor messenger RNA levels in rat dorsal root ganglia after peripheral axotomy.

It is now well established that the expression of peptides in rat primary sensory neurons is dramatically changed in response to peripheral nerve injury. Thus, as first shown by Jessell et al. peripheral axotomy causes a decrease in substance P levels in the dorsal horn of the corresponding spinal cord segments, and this is due to down-regulation of peptide synthesis in dorsal root ganglion neurons. In contrast, other peptides such as vasoactive intestinal polypeptide and peptide histidine isoleucine, galanin and neuropeptide Y are all markedly upregulated in the rat L4 and L5 dorsal root ganglia after sciatic nerve sectioning. The levels of another peptide, cholecystokinin and its messenger RNA are normally very low or undectable in rat primary sensory neurons, but after peripheral axotomy approximately 30% of the ganglion neurons express cholecystokinin messenger RNA. During the last few years a number of peptide receptors have been cloned, and they all belong to the family of G-protein coupled receptors with seven membrane spanning segments, among them the two cholecystokinin receptors cholecystokininA and cholecystokininB. Ghilardi et al. have recently described presence of cholecystokininB binding sites in rat dorsal root ganglia neurons. In the present study we report that the messenger RNA for the cholecystokininB receptor is present at very low levels in normal dorsal root ganglia of the rat, but axotomy causes a very marked increase in the number of sensory neurons of all sizes expressing cholecystokininB receptor messenger RNA, suggesting an increased sensitivity to cholecystokinin for many primary sensory neurons of different modalities after lesion.

Animals↗
PubMed1997

Expression of cholecystokinin messenger RNA in reciprocally-connected auditory thalamus and cortex in the rat.

Cholecystokinin exerts a potent antiepileptic action in mammalian auditory system and undergoes seizure-mediated up-regulation. The present study investigated cholecystokinin messenger RNA expression in the reciprocally-connected auditory thalamus and cortex in the rat. Immunofluorescence in situ hybridization was performed using a 24-base cholecystokinin-messenger RNA oligonucleotide probe. Corticothalamic projection neurons were identified by means of the retrograde fluorescent tracer rhodamine latex microspheres injected into the medial geniculate body. In our experiments, cholecystokinin messenger RNA transcripts were found in about 80% of neurons located within the reciprocally-connected regions of the medial geniculate body and the auditory cortices. These observations provide evidence of cholecystokinin production in the reciprocally-connected regions of the auditory thalamus and cortex, the structures which jointly create the thalamo-corticothalamic circuit which has been implicated in seizure genesis.

Animals↗
PubMed1996

Localization of preprogalanin messenger RNA in rat brain: identification of transcripts in a subpopulation of cerebellar Purkinje cells.

Galanin, a 29 amino acid peptide, is widely distributed throughout both the peripheral and central nervous systems and is thought to be involved in multiple physiological functions including smooth muscle relaxation, stimulation of feeding, blood pressure regulation, control of hormone secretion and modulation of nociception. Galanin has been shown to co-exist with several neurotransmitters throughout the neuroaxis and in some cases to modify their presynaptic and postsynaptic actions. In the present study, the anatomical distribution of preprogalanin messenger RNA in rat brain was examined by in situ hybridization histochemistry using specific 35S-labelled oligonucleotide probes. Neurons expressing preprogalanin messenger RNA were found throughout the brain and were particularly abundant in the hypothalamus. High densities of preprogalanin messenger RNA-positive neurons were found in the anteroventral preoptic, supraoptic, paraventricular and dorsomedial nuclei of the hypothalamus, in the locus coeruleus and in the nucleus of the solitary tract. Moderate densities of preprogalanin messenger RNA-positive cells were apparent in the periventricular and arcuate nuclei of the hypothalamus, in the dorsal raphe and dorsal cochlear nuclei. Low densities of preprogalanin messenger RNA-expressing neurons were observed in the piriform cortex, medial septum and the retrochiasmatic area. These findings are consistent with results of previous in situ localization studies of preprogalanin messenger RNA and also with studies reporting the distribution of galanin-like immunoreactivity in rat brain. A novel finding, however, was the detection of preprogalanin messenger RNA in Purkinje cells in the caudal cerebellar vermis (lobules 6 to 10) and the flocculus and paraflocculus of the lateral hemispheres of the cerebellum. Galanin is presumably co-localized in these cells with GABA, which is normally present in Purkinje cells and possibly with tyrosine hydroxylase, which has recently been detected in a similar subpopulation of cerebellar Purkinje cells in both rat and mouse. Thus, the present study reveals a previously unreported site of galanin gene expression in the cerebellum which represents a novel, putative site of action for galanin to add to its already varied physiological roles.

Animals↗
PubMed1999

An examination of coaxial stacking of helical stems in a pseudoknot motif: the gene 32 messenger RNA pseudoknot of bacteriophage T2.

The RNA pseudoknot located at the 5' end of the gene 32 messenger RNA of bacteriophage T2 contains two A-form helical stems connected by two loops, in an H-type pseudoknot topology. A combination of multidimensional NMR methods and isotope labeling were used to investigate the pseudoknot structure, resulting in a more detailed structural model than provided by earlier homonuclear NMR studies. Of particular significance, the interface between the stacked helical stems within the pseudoknot motif is described in detail. The two stems are stacked in a coaxial manner, with an approximately 18 degrees rotation of stem1 relative to stem2 about an axis that is parallel to the helical axis. This rotation serves to relieve what would otherwise be a relatively close phosphate-phosphate contact at the junction of the two stems, while preserving the stabilizing effects of base stacking. The ability of the NMR data to determine pseudoknot bending was critically assessed. The data were found to be a modestly precise indicator of pseudoknot bending, with the angle between the helical axes of stem1 and stem2 being in the range of 15+/-15 degrees. Pseudoknot models with bend angles within this range are equally consistent with the data, since they differ by only small amounts in the relatively short-range interproton distances from which the structure was derived. The gene 32 messenger RNA pseudoknot was compared with other RNA structures with coaxial or near-coaxial stacked helical stems.

Carbon Isotopes↗
PubMedDate not supplied

[Isolation of interferon messenger RNA and the synthesis of the double-stranded DNA product].

The data on preparation of the fraction of "enriched" interferon messenger RNA of chick fibroblasts by fractionation of biologically active RNA molecules by molecular weight are presented. The conditions of enzymatic synthesis and characteristics of the single- and double-stranded DNA product are described. The ways of producing more highly specific messenger RNA and DNA products are discussed.

Animals↗
PubMed1997

Localization of estrogen receptor protein and estrogen receptor messenger RNA in peripheral autonomic and sensory neurons.

The presence of estrogen receptor protein and estrogen receptor messenger RNA was revealed in peripheral ganglionic neurons of the rat. The pelvic parasympathetic autonomic ganglion and lumbosacral dorsal root sensory ganglia were examined for estrogen receptor-containing neurons because they have known projections to the uterus and uterine cervix. The vagal nodose ganglia were studied for estrogen receptor-containing neurons because they are suspected sources of influence on the uterus. Immunohistochemistry. in situ hybridization histochemistry and retrograde tracing were utilized. Immunoreactivity for estrogen receptors was evident in the nuclei of a subpopulation of neurons in the pelvic ganglia, sixth lumbar and first sacral dorsal root ganglia and nodose ganglia. Some estrogen receptor-positive neurons also contained the retrograde tracer FluoroGold that previously had been injected into the uterus and uterine cervix. Estrogen receptor messenger RNA was also evident in a subpopulation of ganglionic neurons. These data suggest that a certain population of neurons in autonomic and sensory ganglia are capable of synthesizing estrogen receptors and these receptors can serve as binding sites for estrogen. Thus, certain aspects of the structure, function and neurochemistry of some autonomic and sensory neurons may be influenced by the sex steroid estrogen.

Animals↗