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At least 847 records · Page 47Linked to original sources
PubMed1996

Molecular characterization and modular analysis of human MyD88.

MyD88 was first characterized as a myeloid differentiation primary response gene in mice, activated in M1 myeloleukemic cells following interleukin-6 (IL-6) induced growth arrest and terminal differentiation. Analysis of expressed sequence tags (ESTs) from activated dendritic cell libraries led to the indentification of cDNAs encoding the human homolog (hMyD88). The original description of MyD88 as a 243 aa protein may reflect a truncated mouse cDNA since the 2682 nt hMyD88 cDNA predicts a 296 aa cytoplasmic protein. Consistent with this proposal is the detection of a 33 kDa protein in human heart, kidney and liver tissue. The expression pattern of MyD88 is also more widespread than originally believed: a 2.6 kb hMyD88 mRNA species was found to be constitutively expressed in many adult human tissues; in addition MyD88 expression was observed in monocyte, T, B, NK and dendritic cells. The MyD88 protein has a modular structure composed of an N-terminal 'death domain' (DD) similar to the intracellular segments of TNF receptor 1 (TNFR1) and FAS and a C-terminal region related to the signaling domains of vertebrate interleukin-1 receptors (IL-1R) and the Drosophila morphogen Toll. This intriguing structural framework may endow MyD88 with unique signaling capabilities.

Adaptor Proteins, Signal Transducing↗
PubMed1998

Gene expression during adventitious root formation in apple.

A model system for adventitious root formation in woody plants was used to identify transcripts that are up-regulated during this process. 1 mm stem-disks from micropropagated shoots of the apple cultivar Jork 9 can be induced to form roots by treatment with the auxin indole butyric acid (IBA). Stem discs are placed on medium containing IBA for 24 hours and then transferred to IBA-free medium. Root initials become visible after 5-6 days and root elongation occurs within 7-9 days. The first visible cell divisions, which will give rise to the adventitious roots, are detectable 48 hours following IBA treatment. We have used this system to identify transcripts that are induced during adventitious root formation. Two techniques were employed in this analysis: differential messenger RNA display (DDRT) and mRNA representational difference analysis (RDA), a technique that couples PCR and subtractive hybridisation. Using both of these techniques a number of clones have been isolated that exhibit differential expression during auxin induced root formation. Both up-regulated and down-regulated transcripts have been identified. Expression of these genes has initially been verified by reverse northern blot analysis. Northern blot analysis with individual clones has confirmed the expression pattern observed in the reverse northern analysis. Tentative identities of some of the clones have been established by sequencing the partial cDNAs. Among the up-regulated transcripts are clones that share sequence homology with polygalacturonase and MAP kinases. A full-length cDNA for the most abundant up-regulated mRNA, a 2-oxoacid dependent dioxygenase, was characterised. This mRNA is expressed between 24 and 72 hours following IBA treatment of apple stem disks.

Amino Acid Sequence↗
PubMed1998

Use of hydrophilic polymers with microcrystalline cellulose to improve extrusion-spheronization.

Microcrystalline cellulose 19 parts was combined with sodium carboxymethylcellulose, hydroxypropylmethyl cellulose, hydroxypropyl cellulose or polyvinylpyrrolidone one part, either by spray-drying or physical mixing. This combined excipient (20%) mixed with lactose (80%) and water was added to aid formation of pellets by the process of extrusion-spheronization. Spray-dry combined excipient produced pellets with higher yield, better sphericity and improved tolerance to minor variation in the level of water added, compared with the physical mix excipient. Physicochemical testing based on scanning electron microscopy with energy dispersive analysis, differential scanning calorimetry and X-ray diffraction analysis, indicated that the spray-drying with the hydrophilic polymer caused disintegration of the microcrystalline cellulose component into smaller crystallites, favouring its more uniform dispersion throughout the lactose during subsequent processing. The hydroxypropyl cellulose or polyvinylpyrrolidone containing excipients were the most satisfactory of the hydrophilic polymers examined, because they had the least adhesive strength favouring maximum yield of highly spherical pellets.

Calorimetry, Differential Scanning↗
PubMed2004

Transformation of lactulose trihydrate into anhydrous lactulose by fluidized bed drying and its characterization.

Stable anhydrous lactulose was produced from lactulose trihydrate by stepwise heating on a fluidized bed. The processes were performed on stable powder forms. The anhydrous lactulose was characterized by an opaque white appearance, a coarse surface structure with random cracks and indentations, a high degree of crystallization, stability under humid conditions, and by X-ray powder diffraction, differential thermal analysis, and differential thermogravimetry. Those characteristics were different from those of the original trihydrate, which was transparent, had a smooth surface and a higher degree of crystallization, was stable under humid conditions and had different X-ray powder diffraction, differential thermal analysis, and thermogravimetric characteristics. The transformation was enhanced when the inlet temperature was 45-55 degrees C or when the temperature of the fluidized bed was over 40 degrees C. At these cutoff temperatures, both crystalline forms were observed.

Crystallization↗
PubMed1996

Transcriptional responses to tubule challenges.

Adaptation to physiological stimuli often involves changes in gene transcription. Studies of hyperosmolar stress in renal epithelial cells have provided an ideal paradigm for understanding regulation of gene expression. Renal epithelial cells respond very differently to hyperosmolar NaCl and urea and several strategies including cloning based on known biological function, candidate gene analysis, and differential display analysis have successfully identified many genes induced by these hyperosmolar challenges. Hyperosmolar NaCl produces adverse effects on cellular biosynthetic processes and compensatory increases are observed in transcription of transporters, stress proteins, and metabolic enzymes. In contrast, hyperosmolar urea fails to inhibit biosynthetic processes but, nonetheless, initiates a very specific program of gene expression in renal epithelial cells. This program appears to involve a urea sensor/receptor system which activates transcription and translation of the zinc-finger transcription factor Egr-1. This work highlights the concept that rapid analysis of differential gene expression will enable one to define cellular programs of gene expression involving up- and down-regulation of functionally-related gene families.

Animals↗
PubMed2006

Prediction of a key role of motifs binding E2F and NR2F in down-regulation of numerous genes during the development of the mouse hippocampus.

BACKGROUND: We previously demonstrated that gene expression profiles during neuronal differentiation in vitro and hippocampal development in vivo were very similar, due to a conservation of the important second singular value decomposition (SVD) mode (Mode 2) of expression. The conservation of Mode 2 suggests that it reflects a regulatory mechanism conserved between the two systems. In either dataset, the expression vectors of all the genes form two large clusters that differ in the sign of the contribution of Mode 2, which for the majority of them reflects the difference between down- or up-regulation. RESULTS: In the current work, we used a novel approach of analyzing cis-regulation of gene expression in a subspace of a single SVD mode of temporal expression profiles. In the putative upstream regulatory sequences identified by mouse-human homology for all the genes represented in either dataset, we searched for simple features (motifs and pairs of motifs) associated with either sign of the loading of Mode 2. Using a cross-system training-test set approach, we identified E2F binding sites as predictors of down-regulation of gene expression during hippocampal development. NR2F binding sites, for the transcription factors Nr2f/COUP and Hnf4, and also NR2F_SP1 pairs of binding sites, were predictors of down-regulation of expression both during hippocampal development and neuronal differentiation. Analysis of another dataset, from gene profiling of myoblast differentiation in vitro, shows that the conservation of Mode 2 extends to the differentiation of mesenchymal cells. This permitted the identification of two more pairs of motifs, one of which included the CDE/CHR tandem element, as features associated with down-regulation both in the differentiating myoblasts and in the developing hippocampus. Of the features we identified, the E2F and CDE/CHR motifs may be associated with the cycling progenitor cell status, while NR2F may be related to the entry into differentiation along the neuronal pathway. CONCLUSION: Our results constitute the first prediction of an expression pattern from the genomic sequence for the developing mammalian brain, and demonstrate a potential for the analysis of gene regulation in a subspace of a single SVD mode of expression.

Amino Acid Motifs↗
PubMed1996

The prediction of vertebrate promoter regions using differential hexamer frequency analysis.

MOTIVATION: To develop an algorithm utilizing differential hexamer frequency analysis to discriminate promoter from non-promoter regions in vertebrate DNA sequence, without relying upon an extensive database of known transcriptional elements. RESULTS: By determining hexamer frequencies derived from known promoter regions, coding regions and non-coding regions in vertebrates' DNA sequence, and a formula first applied by Claverie and Bougueleret (1986), a discriminant measure was created that compares promoter regions with coding (D1) and non-coding (D2) sequence. The algorithm is able to identify correctly the promoter regions in 18 of 29 loci (62.1%) from an independent test data set. With program options set to identify only one promoter region in the forward strand, there are 11 false-positive predictions in 208 714 nucleotides (one false positive in 18 974 single-stranded bp). With options set to analyze sequence in discrete segments, there is no appreciable improvement in sensitivity, whereas the specificity falls off predictably. It is of particular interest than a search for a peak score (independent of an absolute threshold) is more accurate that a search based upon a fixed scoring threshold. This suggests that the selection of promoter sites may be influenced by the global properties of an entire sequence domain, rather than exclusively upon local characteristics.

Algorithms↗
PubMed2006

Dielectric measurement: error analysis and assessment of uncertainty.

The advantages and limitations of using partial differential analysis to assess the methodological uncertainty associated with the measurement of the dielectric properties of a material are discussed and an alternative pragmatic approach is proposed. It relies on repeat measurements of the dielectric properties of reference liquids and an analysis to estimate random and systematic uncertainties. Examples of measurement uncertainty are provided for well-defined monomolecular materials and for less homogeneous materials at microwave frequencies. All examples relate to measurement with an open-ended coaxial probe but the methodology is not specific to this technique. Examination of the components of uncertainty in the dielectric properties of biological tissue shows that, where the system is free of methodological bias, random fluctuations originating from sampling and natural inhomogeneity dominate the uncertainty budget. In such cases, the mean value of the measured parameter and the standard error of the mean can be taken as a good measure of the true value and its associated uncertainty.

Biometry↗
PubMed1999

Normalization of array hybridization experiments in differential gene expression analysis.

For detecting and confirming differentially expressed genes it is necessary to have a trustworthy reference. So called 'housekeeping genes' are frequently used for this purpose as internal standard. However, if the influence of new experimental conditions is to be analyzed it is not safe to assume a priori that the expression of these genes is not affected. Therefore two synthetic poly(A)-RNAs were generated by PCR and in vitro transcription. They were used as external standards for normalization of northern blots and cDNA arrays where non-regulated genes as internal reference were not available.

Ampicillin Resistance↗
PubMed1994

[The Repertory Grid as a research instrument in inpatient psychotherapy. Possibilities of semantic construct analysis in patients with psychogenic pain].

The Repertory Grid Technique provides a differential analysis of individual constructs ("hypotheses"), by which we structure the perception of our environment. In our study we investigated 35 psychogenic chronic pain patients at beginning and end of inpatient psychotherapy, whether semantic changes occurred in their construct system. We analysed these semantic changes using a categorial system, which was developed by Landfield (1971). This categorial system consists of 22 different categories, at which a rating system (by use of a manual) evaluates the appropriate semantic attachment of constructs of an individual repertory grid to these categories. Our study shows that contrary to our expectation psychogenic pain patients comparatively often described themselves with constructs, which expressed direct social interactions or emotions. However, this finding must be interpreted with some reservation, as our patient group probably is not representative for most of the chronic pain patients, which usually are hardly to motivate for inpatient (respectively outpatient) psychotherapy. Finally we investigated the semantic changes of the construct system during the course of inpatient psychotherapy--here we assume that the conceptualization of own theory guided categories will provide more clinically relevant or perspective results compared to the original category system of Landfield.

Adult↗
PubMed2007

Reduced genetic structure of north Ethiopian cattle revealed by Y-chromosome analysis.

Ethiopia is considered to be a putative migratory corridor for both Near-East Bos taurine and Arabian and Indian B. indicus cattle into East Africa. African pastoralism, which is associated with adaptation to specific habitats and farming systems, has contributed to the composite constitution of Ethiopian cattle. We analyse, for the first time, five Y-chromosome microsatellite markers from seven north Ethiopian cattle populations, using a European Holstein-Friesian population as a reference, to assess the paternal gene pool and to explore the mechanisms behind the genetic structure. Our results reveal that the indicine alleles predominate in the present populations, with only one animal in the Arado carrying the taurine alleles. The north Ethiopian cattle populations with one exception (Abergelle) are characterized by a general low Y-chromosome haplotype diversity, as well as by a reduced interpopulation variance (Phi(ST)=4.0%), which can be a result of strong male-mediated selective sweeps. Population structure revealed by multidimensional-scaling analysis differentiates two populations (Arado and Abergelle) from the rest. Analysis of molecular variance does not lend support to the traditional classification for the populations, which is mainly based on physical characteristics. A network analysis indicates two closely related founding haplotypes accounting for a large proportion (50.0% in Abergelle and 85.0-94.7% in others) of north Ethiopian cattle Y-chromosomes. Our findings point to a common, but limited, paternal origin of the north Ethiopian cattle populations and strong male-mediated gene flow among them. The findings also provide insight into the historical immigration of cattle into East Africa.

Animals↗
PubMed1996

Identification of differentially expressed genes in rat aortic allograft vasculopathy.

Graft vasculopathy is an important complication of long-surviving organ transplants, but its pathogenesis has remained elusive. We investigated rat aortic transplants with vasculopathy, aortic transplants without vasculopathy, and normal aortas for differentially expressed mRNA transcripts to gain further insight into the molecular mechanisms involved. Aortic transplants were performed in allogeneic or syngeneic recipients followed by removal after 1 or 5 months, RNA isolation, and differential display to identify mRNA transcripts the expression of which was modulated in conjunction with the transplant procedure and the development of vasculopathy. Using 80 random primers, 57 differentially displayed polymerase chain reaction products were identified, 18 of which were found in allografts but not in syngeneic grafts or normal vessels, whereas 15 were expressed in normal vessels and syngeneic grafts but not in allografts. Of the differentially displayed amplicons, 13 were successfully reamplified and used as probes for Northern analysis; differential expression was confirmed in 6 instances. DNA sequence analysis of these PCR products revealed identity with the immunoglobulin J chain in 2 instances, the ferritin heavy chain, a sequence related but not identical with Ras, and an established sequence tag recently isolated from a human fetal heart library; 1 sequence was not related to any known gene. To assess whether differential mRNA expression of the J-chain gene, a gene expressed in cells of B lymphocyte lineage, was associated with infiltration of the graft by B lymphocytes, tissue sections were stained with an antibody against the B cell marker CD45RA. Although the number of CD45RA-positive cells was low, there was a significant increase in the number of CD45RA-positive cells in the adventitia and intima of grafts with vasculopathy. Furthermore, immunostaining with anti-ferritin antiserum confirmed the presence of ferritin-positive cells within the inner layer of the graft vessel wall and dispersed in the intima, media, and adventitia. The question remains as to which of these genes are critically relevant in the pathogenesis of graft vasculopathy and whether they serve as targets for therapeutic interventions.

Animals↗
PubMedDate not supplied

Dietary effects of copper and iron deficiency on rat intestine: a differential display proteome analysis.

Copper and iron are cofactors of many metallo-proteins that accomplish vital functions, such as oxygen and electron transport. Specific metabolic pathways have been selected through evolution, although still not fully elucidated, to confine the dangerous reactivity of their free ionic forms. Inadequate supply of both metals can severely affect basic physiological functions. A differential analysis of the rat intestinal proteome evidenced the following dietary copper- and iron-deficiencies, i.e., significant changes in the levels of proteins belonging to different functional classes (glucose and fatty acid metabolism, molecular chaperones, cytoskeleton plasticity, vitamin transporters). The presented results bring new perspectives to understand the role of copper and iron in the metabolic pathways and provide novel diagnostic tools to characterize the effects of subclinical deficiencies of both metals in unbalanced nutritional disorders.

Animals↗
PubMed1993

Generation of structural and functional diversity in furin-like proteins in Drosophila melanogaster by alternative splicing of the Dfur1 gene.

To investigate whether or not alternative splicing might be a mechanism by which in Drosophila melanogaster diversity is generated in endoproteases of the novel eukaryotic family of subtilisin-like proprotein processing enzymes, we determined structural and functional characteristics of the Dfur1 gene. Northern blot analysis revealed Dfur1 transcripts of 7.6, 6.5, 4.5 and 4.0 kb. By comparative nucleotide sequence analysis of Dfur1 genomic and cDNA clones, 10 coding exons were identified and, together with Northern blot analysis using exon-specific probes, evidence was obtained that the four transcripts are generated by alternative splicing and polyadenylation. The apparently complete open reading frames of three Dfur1 cDNAs revealed that these coded for three furin-like proteins, Dfurin1 (892 residues), Dfurin1-CRR (1101 residues) and Dfurin1-X (1269 residues), which possessed common but also unique structural domains. These various isoforms of furin in Drosophila were characterized in gene transfer studies using immunoprecipitation analysis. Differential expression of Dfur1 transcripts was found in Northern blot analysis of RNA from various developmental stages of Drosophila. RNA in situ hybridization experiments revealed that the Dfurin1-X and Dfurin1-CRR isoforms are expressed in non-overlapping sets of tissues during Drosophila embryogenesis. In gene transfer experiments in which the Dfurin1, Dfurin1-CRR and Dfurin1-X proteins were expressed at high levels together with the precursor of the beta A-chain of activin-A, a member of the transforming growth factor beta (TGF beta) superfamily, or the precursor of von Willebrand factor, all three proteins appeared capable of processing these substrates. Our studies indicate that the Dfur1 gene encodes structurally different subtilisin-like proprotein processing enzymes with distinct physiological functions in Drosophila.

Alternative Splicing↗
PubMedDate not supplied

[A technic for the differentiation of 2 metabolically different cell components, in homogenates or tissue sections, from the distribution of lactate dehydrogenase isoenzymes].

The authors report a mathematic-statistical calculation method for a differentiation analysis of LDH-isoenzyme distribution and the results obtained thereby. The necessity for the development of an improved method of analysis for isoenzymes stems from the fact that even homogenates of very small tissue samples (smaller than 1 mg wet weight) show remarkable statistical inhomogenicity in the distribution of the different fractions. This indicates that cells with very different isoenzyme distribution make up the homogenates and that a cellular inhomogenicity exists and that the origin of isoenzymes is non-uniform, respectively. The previously used description of LDH-isoenzymes is thus insufficient, mis-represented and needs a better differentiation. After representation of the theoretical basis for a differentiating method for analysis of two enzymatically different main cell components (A and B), results from embryonic, adult, degenerated, and malignant tissues are presented and discussed. It is shown that in normal and mature tissues there are always two cell components with different isoenzymes next to each other. It seems that via their metabolites there is a correlation between these apparently metabolically different cell components and they seem to be responsible for the biological homoeostasis over the period of life. Presumably the two cell components produce different glycosaminoglycanes (GAG) and collagen types. In connective tissue certain decoupling mechanisms between the cell components seem to be responsible for the self perpetuation and the chronic course of pathological processes.

Cell Fractionation↗
PubMed1984

In vivo metal-ion release from porous titanium-fiber material.

Dense and porous Ti, Ti-alloy, and stainless steel specimens were implanted in canine trabecular bone. After 6 and 12 months the specimens were retrieved together with bone tissue immediately adjacent. The trace metal content in the tissue samples was determined using neutron activation analysis, differential pulse polarography, graphite furnace atomic-absorption spectrophotometry, electron microprobe analysis, and laser microprobe analysis. The results are discussed in view of (i) the release of Ti ions, which is larger for porous than for bulk specimens, (ii) the various artifacts arising in electron microprobe (EMP) and laser microprobe mass analysis (LAMMA) determination of compositional gradients of trace metal-ion content in bone tissue, (iii) the absence of measurable quantities of V in bone tissue, and (iv) the difference in local tissue accumulation between Ti, released from Ti specimens, and Ni, released from stainless steel specimens.

Alloys↗
PubMed1978

Poly(A) polymerase activity during cell cycle and erythropoietic differentiation in erythroleukemic mouse spleen cells.

Poly(A) polymerase activity was studied in lysates of cultured murine erythroleukemic cells (Friend cells). Incorporation of ATP into acid-precipitable products is dependendent on the presence of Mn2+ or Mg2+ and of an RNA primer. The reaction is specific for ATP as the substrate (KM=290 290 micron, it is not inhibited by actinomycin D and only slightly interferred with by ethidium bromide. Cordycepin 5'-triphosphate and sodium pyrophosphate inhibit the enzyme activity. The chain length of the products of the reaction is dependent on the primer concentration and reaches up to 30 nucleotides. Poly(A) polymerase activity is low in resting (G1 phase) cells 75 nmol ATP incorporated/h per 10(6) cells) and increases to a level about twice as high in early S phase of the cell cycle. A possible model for regulation of enzyme activity is discussed. Polymerase activity in the early phase of erythropoietic differentiation of the cells induced by butyric acid does not show any difference in comparison to untreated controls. A decrease in enzyme activity to levels characteristic for cells in G1 phase accompanies shutdown of cell growth in the course of the ongoing differentiation. Analysis of the DNA content of the cells revealed that erythropoietic differentiation of Friend cells induced by butyric acid is characterized by arrest of the cells in G1 phase of the cell cycle. Poly(A) polymerase activity in erythroleukemic cells is thus controlled only by the phase of the cell cycle; it is not affected by changes in gene expression during erythroid differentiation.

Adenosine Triphosphate↗
PubMed2000

T cell immune responses against melanoma and melanocytes in cancer and autoimmunity.

T cell responses specific for melanoma cells and melanocytes appear to be involved in the rejection of melanoma tumors, as well as in the development of autoimmune reactions in patients with Vogt-Koyanagi-Harada disease (VKH), sympathetic ophthalmia, or autoimmune vitiligo. Some of the target antigens for those T cells have been isolated using cDNA expression cloning with melanoma reactive T cells derived from lymphocytes tumor infiltrating (TIL) of patients with melanoma. These include melanocyte specific proteins, such as tyrosinase, TRP1, TRP2, gp100, and MART-1, cancer-testis antigens, and mutated peptides derived from genetic alterations in melanoma cells. Some of the melanoma reactive T cells appear to respond to cryptic or subdominant self epitopes in melanosomal proteins. Modification of those epitopes to increase their immunogenicity by replacement of amino acids at primary anchor residues for peptide/MHC binding, allowed an improvement in immunotherapy for patients with melanoma. Targets for autoreactive T cells against melanocytes in those autoimmune disorders remain to be identified. Isolation of novel target antigens is important for understanding these pathological T cell responses, as well as for developing new diagnostic and treatment methods for these diseases. A variety of techniques, including cDNA expression cloning with T cells, serological analysis of recombinant cDNA expression libraries (SEREX), cDNA subtraction with representational differential analysis (RDA), and serial analysis of gene expression (SAGE) are now being applied to identify novel melanoma/melanocyte antigens recognized by T cells and antibodies.

Autoimmunity↗