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Results for “Gene expression analysis”

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At least 469 records · Page 26Linked to original sources
PubMed2006

Microarray gene expression analysis in atrophying rainbow trout muscle: a unique nonmammalian muscle degradation model.

Muscle atrophy is a physiological response to diverse physiological and pathological conditions that trigger muscle deterioration through specific cellular mechanisms. Despite different signals, the biochemical changes in atrophying muscle share many common cascades. Muscle deterioration as a physiological response to the energetic demands of fish vitellogenesis represents a unique model for studying the mechanisms of muscle degradation in non-mammalian animals. A salmonid microarray, containing 16,006 cDNAs, was used to study the transcriptome response to atrophy of fast-switch muscles from gravid rainbow trout compared with sterile fish. Eighty-two unique transcripts were upregulated and 120 transcripts were downregulated in atrophying muscles. Transcripts having gene ontology identifiers were grouped according to their functions. Muscle deterioration was associated with elevated expression of genes involved in the catheptic and collagenase proteolytic pathways; the aerobic production, buffering, and utilization of ATP; and growth arrest; whereas atrophying muscle showed downregulation of genes encoding a serine proteinase inhibitor, enzymes of anaerobic respiration, muscle proteins as well as genes required for RNA and protein biosynthesis/processing. Therefore, gene transcription of the trout muscle atrophy changed in a manner similar to mammalian muscle atrophy. These changes result in an arrest of normal cell growth, protein degradation, and decreased glycolytic cellular respiration that is characteristic of the fast-switch muscle. For the first time, other changes/mechanisms unique to fish were discussed including genes associated with muscle atrophy.

Animals↗
PubMed2005

Gene expression analysis of the function of the male-specific lethal complex in Drosophila.

Dosage compensation refers to the equal expression of X-linked genes despite the difference in copy number between the two sexes. The male-specific lethal (MSL) complex is concentrated on the X chromosome in males. A gene expression assay for embryos was developed to examine the function of this complex. In mutant male embryos without either the MSL complex or MOF histone acetylase, dosage compensation is retained but autosomal expression is increased. Dosage compensation is lost in the double-mutant embryos. In embryos in which the MSL complex and MOF are targeted to the X chromosomes in females, the results are consistent with previous surveys showing that in general the X expression remains unchanged, but autosomal expression is reduced. Mutations in the ISWI chromatin-remodeling component cause increases specifically of X-linked genes in males. Thus, the function of the MSL complex in conjunction with ISWI is postulated to override the effect on gene expression of high histone acetylation on the male X. The basic determinant of dosage compensation is suggested to be an inverse dosage effect produced by an imbalance of transcription factors on the X vs. the autosomes. The sequestration of the MSL complex to the male X may have evolved to counteract a similar effect on the autosomes and to prevent an overexpression of the X chromosome in males that would otherwise occur due to the high levels of histone acetylation.

Acetyltransferases↗
PubMed2000

Quantitative gene expression analysis in microdissected archival tissue by real-time RT-PCR.

Laser-assisted microdissection is a powerful tool for the analysis of morphologically defined cell populations. We report here that the combination of microdissection and real-time RT-PCR technologies together with an optimized RNA microscale extraction procedure allows the reproducible determination of gene expression levels in archival formalin-fixed, paraffin-embedded tissue samples.

Dissection↗
PubMed2006

Gene expression analysis for the prediction of recurrence in patients with primary Ta urothelial cell carcinoma.

OBJECTIVES: The individual recurrence-free period after primary surgery of patients with Ta urothelial cell carcinoma (UCC) cannot be predicted accurately. This study aims at discriminating between patients with primary Ta UCC and long or short recurrence-free periods. METHODS: We investigated mRNA expression of 23 genes in 44 primary Ta tumours (23 and 21 tumours were from patients with long [>or=4 yr] or short [<or=2 yr] recurrence-free periods, respectively), using real-time quantitative polymerase chain reaction. The genes were selected from previously published studies and showed a relationship with tumour recurrence in patients with UCC. RESULTS: Differential mRNA expression between the two patient groups indicated statistical significance only for the gene survivin (p=0.0011). Its recurrence predictive value could not be increased by a combination with any of the other genes. Comparison of the receiver operating characteristic curves for survivin expression between patients with long or short recurrence-free intervals revealed an area under the curve of 0.79 (95%CI, 0.65-0.92). Using the median expression (0.84) as cut-off level, survivin identified 71.4% (95%CI, 47.8-88.7) and 69.6% (95%CI, 47.1-86.8) of the patients with long or short recurrence-free periods, respectively. CONCLUSIONS: Our study identifies survivin as the most promising candidate to distinguish between patients with primary Ta UCC and long or short recurrence-free intervals. Therefore, survivin mRNA expression analysis might help the urologist to individualise patient treatment and prevent unnecessary cystoscopies in a subgroup of these patients.

Adult↗
PubMed2003

cDNA cloning and gene expression analysis of human myo-inositol 1-phosphate synthase.

myo-Inositol 1-phosphate synthase (EC 5.5.1.4) (IPS) is a key enzyme in myo-inositol biosynthesis pathway. This study describes the molecular cloning of the full length human myo-inositol 1-phosphate synthase (hIPS) cDNA, tissue distribution of its mRNA and characterizes its gene expression in cultured HepG2 cells. Human testis, ovary, heart, placenta, and pancreas express relatively high level of hIPS mRNA, while blood leukocyte, thymus, skeletal muscle, and colon express low or marginal amount of the mRNA. In the presence of glucose, hIPS mRNA level increases 2- to 4-fold in HepG2 cells. hIPS mRNA is also up-regulated 2- to 3-fold by 2.5 microM lovastain. This up-regulation is prevented by mevalonic acid, farnesol, and geranylgeraniol, suggesting a G-protein mediated signal transduction mechanism in the regulation of hIPS gene expression. hIPS mRNA expression is 50% suppressed by 10mM lithium ion in these cells. Neither 5mM myo-inositol nor the three hormones: estrogen, thyroid hormone, and insulin altered hIPS mRNA expression in these cells.

Amino Acid Sequence↗
PubMed2006

Microfluidic isolation of leukocytes from whole blood for phenotype and gene expression analysis.

Technologies that enable the isolation of cell subtypes from small samples of complex populations will greatly facilitate the implementation of proteomics and genomics to human diseases. Transcriptome analysis of blood requires the depletion of contaminating erythrocytes. We report an automated microfluidic device to rapidly deplete erythrocytes from whole blood via deionized water lysis and to collect enriched leukocytes for phenotype and genomic analyses. Starting with blood from healthy subjects, we demonstrate the utility of this microfluidic cassette and lysis protocol to prepare unstimulated leukocytes, and leukocytes stimulated ex vivo with Staphylococcal enterotoxin B, which mimics some of the cellular effects seen in patients with severe bacterial infections. Microarrays are used to assess the global gene expression response to enterotoxin B. The results demonstrate that this system can isolate unactivated leukocytes from small blood samples without any significant loss, which permits more information to be obtained from subsequent analysis, and will be readily applicable to clinical settings.

Cell Separation↗
PubMed2006

A comparative gene-expression analysis of CD34+ hematopoietic stem and progenitor cells grown in static and stirred culture systems.

Static and stirred culture systems are widely used to expand hematopoietic cells, but differential culture performances are observed between these systems. We hypothesize that these differential culture outcomes are caused by the physiological responses of CD34(+) hematopoietic stem and progenitor cells (HSPCs) to the different physical microenvironments created in these culture devices. To understand the genetic changes provoked by culture microenvironments, the gene expression profiling of CD34(+) HSPCs grown in static and stirred culture systems was compared using SMART-PCR and cDNA arrays. The results revealed that 103 and 99 genes were significantly expressed in CD34(+) cells from static and stirred systems, respectively. Of those, 91 have similar levels of expression, while 12 show differential transcription levels. These differentially expressed genes are mainly involved in anti-oxidation, DNA repair, apoptosis, and chemotactic activity. A quantitative molecular understanding of the influences of growth microenvironments on transcriptional events in CD34(+) HSPCs should give new insights into optimizing culture strategies to produce hematopoietic cells.

Antigens, CD34↗
PubMed1998

Probing lymphocyte biology by genomic-scale gene expression analysis.

The identity and abundance of mRNA species within a cell dictate, to a large extent, the biological potential of that cell. Although posttranscriptional mechanisms modify protein expression in critical ways, cellular differentiation requires key changes in gene transcription, as evidenced by the potent phenotypes that result from disruption of transcription factor genes in mice. It is now possible to assess the mRNA profile of a cell globally using recently developed genomics techniques. This review focuses on the potential of cDNA microarrays to define gene expression in lymphoid cells, a field which is in its infancy. Examples of cellular activation genes and cytokine inducible genes discovered using this technology are presented but these represent only a taste of the fruit that this new technology will ultimately bear. Gene expression profiles should provide essential new insights into lymphocyte differentiation and activation, the pathogenesis of immune disorders, and the molecular abnormalities in lymphoid malignancies.

DNA, Complementary↗
PubMed2005

Global gene expression analysis of keloid fibroblasts in response to electron beam irradiation reveals the involvement of interleukin-6 pathway.

Keloid is a dermal fibroproliferative lesion of unknown etiology that commonly recurs after surgical excision. Post-operative adjuvant electron beam (EB) irradiation has been successfully used to reduce keloid recurrences. To provide new insights into the molecular mechanism behind the effect of EB irradiation, we used a cDNA microarray screening of more than 5000 genes to assess early changes in gene expression between EB-irradiated and non-irradiated keloid and non-lesional dermal fibroblasts. Primary fibroblast cultures from keloid and associated non-lesional dermis obtained from five patients were exposed to 15 Gy EB irradiation and analyzed after 15 min incubation. Early response to EB irradiation showed that 96 (1.8%) genes were modulated 2-fold or more in keloid fibroblasts. Upregulated genes accounted for 29.2% (28 genes), whereas downregulated genes comprised 70.8% (68 genes), indicating a silencing of many genes in keloid fibroblasts after EB irradiation. Many of the downregulated genes play roles in the enhancement of cell proliferation and extracellular matrix production, whereas several of the upregulated genes involves in the promotion of apoptosis and extracellular matrix (ECM) degradation. Using emerging bioinformatic tools and further corroboration, the interleukin 6 (IL-6) signaling pathway was found to be mainly involved in EB irradiation response. We also showed co-expression of IL-6 and its specific receptor (IL-6Ralpha) in keloid fibroblasts that points to the existence of an IL-6 autocrine loop in these cells. These results suggested that at the molecular level, EB irradiation might hinder keloid formation by regularizing disturbances in the homeostatic equilibrium between inducer and inhibitor activities in the matrix system most likely through the IL-6 pathway. Our study provides clues for the molecular mechanism(s) behind the beneficial effect of EB irradiation in reducing keloid recurrences and may help develop alternative strategies for the therapy and prophylaxis of this lesion.

Adult↗
PubMed2005

Gene expression analysis by cDNA-AFLP highlights a set of new signaling networks and translational control during seed dormancy breaking in Nicotiana plumbaginifolia.

Seed dormancy in Nicotiana plumbaginifolia is characterized by an abscisic acid accumulation linked to a pronounced germination delay. Dormancy can be released by 1 year after-ripening treatment. Using a cDNA-amplified fragment length polymorphism (cDNA-AFLP) approach we compared the gene expression patterns of dormant and after-ripened seeds, air-dry or during one day imbibition and analyzed 15,000 cDNA fragments. Among them 1020 were found to be differentially regulated by dormancy. Of 412 sequenced cDNA fragments, 83 were assigned to a known function by search similarities to public databases. The functional categories of the identified dormancy maintenance and breaking responsive genes, give evidence that after-ripening turns in the air-dry seed to a new developmental program that modulates, at the RNA level, components of translational control, signaling networks, transcriptional control and regulated proteolysis.

Blotting, Northern↗
PubMed2004

Microdissected region-specific gene expression analysis with methacarn-fixed, paraffin-embedded tissues by real-time RT-PCR.

We have previously shown methacarn to be a versatile fixative for analysis of proteins, DNA, and RNA in paraffin-embedded tissues (PETs). In this study we analyzed its suitability for quantitative mRNA expression analysis of microdissected PET specimens using a real-time RT-PCR technique. Fidelity of expression in the methacarn-fixed PET sections, with reference to dose-dependent induction of cytochrome P450 2B1 in the phenobarbital-treated rat liver, was high in comparison with the unfixed frozen tissue case, even after hematoxylin staining. RNA yield from methacarn-fixed PET sections was equivalent to that in unfixed cryosections and was also not significantly affected by hematoxylin staining. Correlations between the expression levels of target genes and input amounts of extracted RNA in the range of 1-1000 pg were very high (correlation coefficients >0.98), the regression curves being similar to those with unfixed cryosections. Although cell numbers should be optimized for each target gene/tissue, >/=200 cells were necessary for accurate measurement in 10-microm-thick rat liver sections judging from the variation of measured value in small microdissected areas. These results indicate high performance with methacarn, close to that of unfixed tissues, regarding quantitative expression analysis of mRNAs in microdissected PET-specimens.

Acetic Acid↗
PubMed2000

A polymorphism in the mouse crg-2/IP-10 gene complicates chemokine gene expression analysis using a commercial ribonuclease protection assay.

The use of multiprobe RPAs is becoming an increasingly popular method for the detection and quantitation of RNA levels in cells and tissues. Here we report that due to a polymorphism in the 3'-noncoding region of the mouse Crg-2/IP-10 gene, the mCK-5 chemokine probe set available from Pharmingen can yield aberrant signal patterns with RNA samples from BALB/c, MRL and possibly other mouse strains that may lead to false conclusions regarding expression of the Crg-2/IP-10 and MCP-1 genes.

Animals↗
PubMed2004

Gene expression analysis of peripheral T cells in a subgroup of common variable immunodeficiency shows predominance of CCR7(-) effector-memory T cells.

Common variable immunodeficiency (CVID) represents a heterogeneous group of antibody deficiency syndromes, characterized by defective antibody production in which T cell deficiency may play a pathogenic role. A subgroup of CVID patients has impaired in vitro T cell proliferation. Using microarray analyses of T cells from these patients, we found a gene expression pattern different from healthy controls and patients with X-linked agammaglobulinaemia. The profile of the differentially expressed genes suggests enhanced cytotoxic effector functions, antigen experienced or chronically activated T cells and a predominance of CCR7(-) T cells. Further experiments using flow cytometry revealed a striking predominance of CCR7(-) T cells in a subgroup of CVID patients, and an association with impaired T cell proliferation. Our observations indicate that a predominance of CCR7(-) T cells with effector-memory cell features and with reduced proliferative capacity may characterize a subgroup of CVID.

Adult↗
PubMed2006

Gene expression analysis in mice liver on hepatocarcinogenesis by flumequine.

mRNA expression profiles in the liver from mice treated with flumequine (FL) were analyzed in order to elucidate the mechanism of its tumor-promoting effect. The liver from a C3H/He mouse that received a diet containing 4,000 ppm of FL for 4 weeks was examined by cDNA microarray in comparison with an untreated mouse. Furthermore, to obtain a more comprehensive sequence, time-course changes in selected genes were determined by real-time RT-PCR. Microarray analysis revealed 15 upregulated and 9 downregulated genes in an FL-treated mouse. The upregulated genes included signal transducers and cell cycle regulators. In addition, the levels of stress response genes, particularly glutathione S-transferase (GST) alpha and GSTmu, were very high, indicating the generation of oxidative stress. On the other hand, the downregulated genes included phase I metabolic enzymes, such as cytochrome P450 (CYPs) enzymes, and apoptosis-associated proteins. These changes were confirmed by quantitative RT-PCR and were generally consistent with each other. Time-course observations revealed consistent results, particularly with regard to GSTalpha, GSTmu, ERK5, and CYP2E1. In addition, the expression of 8-oxoguanine DNA glycosylase 1 (OGG1) was increased in a time-dependent manner. These results suggest the possibility that responses against oxidative stress may play a major role in hepatocarcinogenesis by FL in mice.

Animals↗
PubMed2003

Gene expression analysis of energy metabolism mutants of Desulfovibrio vulgaris Hildenborough indicates an important role for alcohol dehydrogenase.

Comparison of the proteomes of the wild-type and Fe-only hydrogenase mutant strains of Desulfovibrio vulgaris Hildenborough, grown in lactate-sulfate (LS) medium, indicated the near absence of open reading frame 2977 (ORF2977)-coded alcohol dehydrogenase in the hyd mutant. Hybridization of labeled cDNA to a macroarray of 145 PCR-amplified D. vulgaris genes encoding proteins active in energy metabolism indicated that the adh gene was among the most highly expressed in wild-type cells grown in LS medium. Relative to the wild type, expression of the adh gene was strongly downregulated in the hyd mutant, in agreement with the proteomic data. Expression was upregulated in ethanol-grown wild-type cells. An adh mutant was constructed and found to be incapable of growth in media in which ethanol was both the carbon source and electron donor for sulfate reduction or was only the carbon source, with hydrogen serving as electron donor. The hyd mutant also grew poorly on ethanol, in agreement with its low level of adh gene expression. The adh mutant grew to a lower final cell density on LS medium than the wild type. These results, as well as the high level of expression of adh in wild-type cells on media in which lactate, pyruvate, formate, or hydrogen served as the sole electron donor for sulfate reduction, indicate that ORF2977 Adh contributes to the energy metabolism of D. vulgaris under a wide variety of metabolic conditions. A hydrogen cycling mechanism is proposed in which protons and electrons originating from cytoplasmic ethanol oxidation by ORF2977 Adh are converted to hydrogen or hydrogen equivalents, possibly by a putative H(2)-heterodisulfide oxidoreductase complex, which is then oxidized by periplasmic Fe-only hydrogenase to generate a proton gradient.

Alcohol Dehydrogenase↗
PubMed2006

Genetic linkage map and expression analysis of genes expressed in the lamellae of the edible basidiomycete Pleurotus ostreatus.

Pleurotus ostreatus is an industrially cultivated basidiomycete with nutritional and environmental applications. Its genome contains 35 Mbp organized in 11 chromosomes. There is currently available a genetic linkage map based predominantly on anonymous molecular markers complemented with the mapping of QTLs controlling growth rate and industrial productivity. To increase the saturation of the existing linkage maps, we have identified and mapped 82 genes expressed in the lamellae. Their manual annotation revealed that 34.1% of the lamellae-expressed and 71.5% of the lamellae-specific genes correspond to previously unknown sequences or to hypothetical proteins without a clearly established function. Furthermore, the expression pattern of some genes provides an experimental basis for studying gene regulation during the change from vegetative to reproductive growth. Finally, the identification of various differentially regulated genes involved in protein metabolism suggests the relevance of these processes in fruit body formation and maturation.

Chromosome Mapping↗
PubMed1999

Alginate-poly-L-lysine microcapsule biocompatibility: a novel RT-PCR method for cytokine gene expression analysis in pericapsular infiltrates.

Transplantation of microencapsulated islets of Langerhans is impaired by a pericapsular host reaction that eventually induces graft failure. We are studying the role of cytokines in the pathogenesis of this reaction, using the model of alginate-polylysine microcapsule implantation in rat epididymal fat pads. The objectives were: (1) to develop a method to measure, by semiquantitative PCR, TGF-beta1 gene expression in fat pad pericapsular infiltrates, and (2) to use this method to evaluate TGF-beta1 gene expression 14 days after microcapsule implantation. TGF-beta1 mRNA level was significantly higher in pericapsular infiltrate cells than in nonimplanted tissue cells and saline-injected tissue cells (p < 0.0001 and p < 0.01, respectively). There was no significant difference between the TGF-beta1 mRNA levels of the two types of controls (p = 0.0945). These results suggest that TGF-beta1 plays a role in the pathogenesis of the pericapsular reaction. The method developed can be used to study the role of other fibrogenic cytokines potentially involved. This will shed light on the mechanisms underlying the pericapsular reaction and will serve as a basis for the development of strategies to control this reaction.

Adipose Tissue↗