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Protein Expression Patterns (protein + expression_pattern)
Selected AbstractsProtein expression pattern of P,glycoprotein along the gastrointestinal tract of the yucatan micropigJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 1 2004Huadong Tang Abstract The purpose of this study is to characterize the distribution pattern of P,gp protein levels along the entire GI tract in the Yucatan micropig, which is being developed as a model for human drug bioavailability. Small and large intestines were freshly obtained and divided into about 37 segments and 10 segments, respectively (ca., 1 foot/segment). Epithelial cells from the small intestine were obtained by an elution method; whereas, a scraping method was applied to the large intestine. Total cellular protein was isolated from the epithelial cells. Western blot analysis using P,gp antibody showed that the amount of P,gp protein increased distally from the duodenum to the ileum over approximately a 10,fold range. P,gp protein in the large intestine was present at a higher level in the central portion, but the absolute amount was much less than what was found in the small intestine. © 2004 Wiley Periodicals, Inc. J Biochem Mol Toxicol 18:18,22, 2004; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20001 [source] Ontogeny of tyrosine hydroxylase mRNA expression in mid- and forebrain: Neuromeric pattern and novel positive regionsDEVELOPMENTAL DYNAMICS, Issue 3 2005Faustino Marín Abstract Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the synthesis of catecholamines and, thus, critical in determining the catecholaminergic phenotype. In this study, we have examined the expression of TH mRNA by in situ hybridization in the embryonic mouse forebrain and midbrain and have mapped its localization according to the neuromeric pattern. We find that early in embryonic development, 10 to 12 days post coitum (dpc), TH mRNA is expressed in ample continuous regions of the neuroepithelium, extending across several neuromeres. However, from 12.5 dpc onward, the expression becomes restricted to discrete regions, which correspond to the dopaminergic nuclei (A8 to A15). In addition to these nuclei previously described, TH mRNA is also observed in regions that do not express this enzyme according to immunohistochemical studies. This difference in relation to protein expression pattern is consequent with the known posttranscriptional regulation of TH expression. The most representative example of a novel positive region is the conspicuous mRNA expression in both medial and lateral ganglionic eminences. This result agrees with reports describing the capacity of striatal stem cells (that is, located at the lateral ganglionic eminence) to become dopaminergic in vitro. Other regions include the isthmic mantle layer and the early floor plate of the midbrain,caudal forebrain. On the whole, the expression map we have obtained opens new perspectives for evolutionary/comparative studies, as well as for therapeutic approaches looking for potentially dopaminergic cells. Developmental Dynamics 234:709,717, 2005. © 2005 Wiley-Liss, Inc. [source] HIF-1, protein expression is associated with the environmental inflammatory reaction in Barrett's metaplasiaDISEASES OF THE ESOPHAGUS, Issue 8 2009F. C. Ling SUMMARY The oxygen-regulated transcription factor subunit hypoxia inducible factor-1, (HIF-1,) is involved in angiogenesis, energy metabolism, cell survival, and inflammation. We examined the protein expression of HIF-1, within the progression of Barrett's sequence as well as the type and degree of the environmental inflammatory reaction. Squamous epithelium (SE), metaplastic, low- and high-grade dysplastic lesions, and tumor tissue of 57 resection specimens from patients with Barrett's adenocarcinoma were immunohistochemically analyzed. Active and chronic inflammatory reactions were classified according to the Updated Sydney System. HIF-1, protein expression increased significantly from SE to Barrett's metaplasia (BM) (P < 0.0001). From metaplasia through low- and high-grade dysplasia to cancer, no further increase could be detected. Active and chronic inflammation were also significantly different between SE and BM (P < 0.0001) but not during further progression in the sequence. HIF-1, protein expression did not correlate with histopathologic parameters or survival. HIF-1, protein expression pattern resembles the active and chronic environmental inflammatory reaction. All were significantly increased in metaplasia compared to SE without further change in tumor development. HIF-1, protein expression appears to be associated with inflammatory processes in the development of BM. [source] Sample pooling in 2-D gel electrophoresis: A new approach to reduce nonspecific expression backgroundELECTROPHORESIS, Issue 22 2006Marc Weinkauf Abstract Protein expression alterations unrelated to an investigated phenotype are accumulated in most cell line models during establishment. Performing a whole proteome screening of lymphoma cell lines, we established a method to reduce the influence of protein expression unrelated to the distinct investigated phenotype. In 2-D PAGE, the comprehensive analysis of a large number of protein spots would be simplified by pooling cell line samples of the investigated phenotype. Applying this pooling approach, unrelated alterations of single samples are ,muted' by dilution. Analysing two different lymphoma subtypes (follicular and mantle cell lymphoma) by this method, spots originating in only single cell lines were reduced by 72% (650/900), whereas even modestly altered expression of protein spots detected in all lines were reliably detected in the pooled protein gels. We conclude that our pooling approach is a preferable approach to reliably detect a common protein expression pattern and may even allow proteomic analysis of clinical samples with limited amounts of sample material, even with minimal cell numbers as low as 1×106. [source] Silencing of an abdominal Hox gene during early development is correlated with limb development in a crustacean trunkEVOLUTION AND DEVELOPMENT, Issue 2 2010Cheryl C. Hsia SUMMARY We tested whether Artemia abd-A could repress limbs in Drosophila embryos, and found that although abd-A transcripts were produced, ABD-A protein was not. Similarly, developing Artemia epidermal cells showed expression of abd-A transcripts without accumulation of ABD-A protein. This finding in Artemia reveals a new variation in Hox gene function that is associated with morphological evolution. In this case, a HOX protein expression pattern is completely absent during early development, although the HOX protein is expressed at later stages in the central nervous system in a "homeotic-like" pattern. The combination of an absence of ABD-A protein expression in the Artemia limb primordia and the weak repressive function of Artemia UBX protein on the limb-promoting gene Dll are likely to be two reasons why homonomous limbs develop throughout the entire Artemia trunk. [source] Proteomic analysis of Arabidopsis halleri shoots in response to the heavy metals cadmium and zinc and rhizosphere microorganismsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 21 2009Silvia Farinati Abstract Arabidopsis halleri has the rare ability to colonize heavy metal-polluted sites and is an emerging model for research on adaptation and metal hyperaccumulation. The aim of this study was to analyze the effect of plant,microbe interaction on the accumulation of cadmium (Cd) and zinc (Zn) in shoots of an ecotype of A. halleri grown in heavy metal-contaminated soil and to compare the shoot proteome of plants grown solely in the presence of Cd and Zn or in the presence of these two metals and the autochthonous soil rhizosphere-derived microorganisms. The results of this analysis emphasized the role of plant,microbe interaction in shoot metal accumulation. Differences in protein expression pattern, identified by a proteomic approach involving 2-DE and MS, indicated a general upregulation of photosynthesis-related proteins in plants exposed to metals and to metals plus microorganisms, suggesting that metal accumulation in shoots is an energy-demanding process. The analysis also showed that proteins involved in plant defense mechanisms were downregulated indicating that heavy metals accumulation in leaves supplies a protection system and highlights a cross-talk between heavy metal signaling and defense signaling. [source] Neuroproteomics and its applications in research on nicotine and other drugs of abusePROTEOMICS - CLINICAL APPLICATIONS, Issue 11 2007Ming D. Li Dr. Abstract The rapidly growing field of neuroproteomics is able to track changes in protein expression and protein modifications underlying various physiological conditions, including the neural diseases related to drug addiction. Thus, it presents great promise in characterizing protein function, biochemical pathways, and networks to understand the mechanisms underlying drug dependence. In this article, we first provide an overview of proteomics technologies and bioinformatics tools available to analyze proteomics data. Then we summarize the recent applications of proteomics to profile the protein expression pattern in animal or human brain tissues after the administration of nicotine, alcohol, amphetamine, butorphanol, cocaine, and morphine. By comparing the protein expression profiles in response to chronic nicotine exposure with those appearing in response to treatment with other drugs of abuse, we identified three biological processes that appears to be regulated by multiple drugs of abuse: energy metabolism, oxidative stress response, and protein degradation and modification. Such similarity indicates that despite the obvious differences among their chemical properties and the receptors with which they interact, different substances of abuse may cause some similar changes in cellular activities and biological processes in neurons. [source] Cell volume and rate of proliferation, but not protein expression pattern, distinguish pup/intimal smooth muscle cells from subcultured adult smooth muscle cellsCELL PROLIFERATION, Issue 5 2001E. McKilligin Smooth muscle cells from neonatal rats and from injured blood vessels grow with a characteristic cobblestone morphology that distinguishes them from adult smooth muscle cells. This has led to the proposition that there are two distinct types of smooth muscle cells with different proliferative capacity. Here we systematically compare the properties of subcultured adult smooth muscle cells in culture and clonal lines of cobblestone smooth muscle cells from both neonatal rats and injured vessels. The cobblestone smooth muscle cells have a significantly smaller average cell volume, estimated using two different flow cytometry measurements. However, the two types of smooth muscle cells have indistinguishable protein expression patterns when the levels of more than 20 different proteins (including cytoskeletal proteins, matrix proteins, cytokines, cytokine receptors, adhesion molecules and enzymes) are measured by quantitative immunofluorescence. Furthermore, in contrast to previous observations, we demonstrate that both types of smooth muscle cells secrete a powerful mitogenic activity. The higher cell density achieved by the cobblestone smooth muscle cells in culture was responsible for the earlier reports that this mitogenic activity was secreted only by cobblestone smooth muscle cells. We conclude that many of the differences seen between cobblestone smooth muscle cells and adult smooth muscle cells in vitro (proliferation rate, morphology, protein expression pattern, secretion of mitogenic activity) could be attributable to a stable difference in the median cell volume of the cultures. [source] Medaka Oct4 is expressed during early embryo development, and in primordial germ cells and adult gonadsDEVELOPMENTAL DYNAMICS, Issue 2 2010Ana V. Sánchez-Sánchez Abstract Oct4 is a crucial transcription factor for controlling pluripotency in embryonic stem cells and the epiblast of mouse embryos. We have characterized the expression pattern of medaka (Oryzias latipes) Ol-Oct4 during embryonic development and in the adult gonads. Genomic analysis showed that Ol-Oct4 is the ortholog of zebrafish spg/pou2. However, their expression patterns are not the same, suggesting that Oct4 may play different roles in zebrafish and medaka. Using specific antibodies for the Ol-Oct4 protein, we showed that Ol-Oct4 is also expressed in primordial germ cells, in the spermatogonia (male germ stem cells), and during different stages of oocyte development. These results suggest that Ol-Oct4 plays a post-embryonic role in the maturing gonads and gametes. The Ol-Oct4 mRNA and protein expression patterns are similar to those of mammalian Oct4 and introduce medaka fish as a valid model for the functional and evolutionary study of pluripotency genes in vivo. Developmental Dynamics 239:672,679, 2010. © 2009 Wiley-Liss, Inc. [source] Platelet-derived growth factors in the developing avian heart and maturating coronary vasculatureDEVELOPMENTAL DYNAMICS, Issue 4 2005Nynke M.S. Abstract Platelet-derived growth factors (PDGFs) are important in embryonic development. To elucidate their role in avian heart and coronary development, we investigated protein expression patterns of PDGF-A, PDGF-B, and the receptors PDGFR-, and PDGFR-, using immunohistochemistry on sections of pro-epicardial quail,chicken chimeras of Hamburger and Hamilton (HH) 28,HH35. PDGF-A and PDGFR-, were expressed in the atrial septum, sinus venosus, and throughout the myocardium, with PDGFR-, retreating to the trabeculae at later stages. Additionally, PDGF-A and PDGFR-, were present in outflow tract cushion mesenchyme and myocardium, respectively. Small cardiac nerves and (sub)epicardial cells expressed PDGF-B and PDGFR-,. Furthermore, endothelial cells expressed PDGF-B, while vascular smooth muscle cells and interstitial epicardium-derived cells expressed PDGFR-,, indicating a role in coronary maturation. PDGF-B is also present in ventricular septal development, in the absence of any PDGFR. Epicardium-derived cells in the atrioventricular cushions expressed PDGFR-,. We conclude that all four proteins are involved in myocardial development, whereas PDGF-B and PDGFR-, are specifically important in coronary maturation. Developmental Dynamics 233:1579,1588, 2005. © 2005 Wiley-Liss, Inc. [source] Systemic RNAi of the cockroach vitellogenin receptor results in a phenotype similar to that of the Drosophila yolkless mutantFEBS JOURNAL, Issue 2 2006Laura Ciudad During vitellogenesis, one of the most tightly regulated processes in oviparous reproduction, vitellogenins are incorporated into the oocyte through vitellogenin receptor (VgR)-mediated endocytosis. In this paper, we report the cloning of the VgR cDNA from Blattella germanica, as well as the first functional analysis of VgR following an RNA interference (RNAi) approach. We characterized the VgR, VgR mRNA and protein expression patterns in pre-adult and adult stages of this cockroach, as well as VgR immunolocalization in ovarioles, belonging to the panoistic type. We then specifically disrupted VgR gene function using RNAi techniques. Knockdown of VgR expression led to a phenotype characterized by low yolk content in the ovary and high vitellogenin concentration in the haemolymph. This phenotype is equivalent to that of the yolkless mutant of Drosophila melanogaster, which have the yl (VgR) gene disrupted. The results additionally open the perspective that development genes can be functionally analyzed via systemic RNAi in this basal species. [source] Modulation of white adipose tissue proteome by aging and calorie restrictionAGING CELL, Issue 5 2010Adamo Valle Summary Aging is associated with an accrual of body fat, progressive development of insulin resistance and other obesity comorbidities that contribute to decrease life span. Caloric restriction (CR), which primarily affects energy stores in adipose tissue, is known to extend life span and retard the aging process in animal models. In this study, a proteomic approach combining 2-DE and MS was used to identify proteins modulated by aging and CR in rat white adipose tissue proteome. Proteomic analysis revealed 133 differentially expressed spots, 57 of which were unambiguously identified by MS. Although CR opposed part of the age-associated protein expression patterns, many effects of CR were on proteins unaltered by age, suggesting that the effects of CR on adipose tissue are only weakly related to those of aging. Particularly, CR and aging altered glucose, intermediate and lipid metabolism, with CR enhancing the expression of enzymes involved in oxalacetate and NADPH production, lipid biosynthesis and lipolysis. Consistently, insulin-, and ,3-adrenergic receptors were also increased by CR, which denotes improved sensitivity to lipogenic/lipolytic stimuli. Other beneficial outcomes of CR were an improvement in oxidative stress, preventing the age-associated decrease in several antioxidant enzymes. Proteins involved in cytoskeleton, iron storage, energy metabolism and several proteins with novel or unknown functions in adipose tissue were also modulated by age and/or CR. Such orchestrated changes in expression of multiple proteins provide insights into the mechanism underlying CR effects, ultimately allowing the discovery of new markers of aging and targets for the development of CR-mimetics. [source] Characterization of childhood precursor T-lymphoblastic lymphoma by immunophenotyping and fluorescent in situ hybridization: A report from the Children's Oncology GroupPEDIATRIC BLOOD & CANCER, Issue 4 2008Kristi J. Smock MD Abstract Background T-lymphoblastic lymphoma (T-LBL) accounts for 25,30% of childhood non-Hodgkin's lymphoma and is closely related to T-lymphoblastic leukemia (T-ALL). Recently, we demonstrated distinct differences in gene expression between childhood T-LBL and T-ALL, but molecular pathogenesis and relevant protein expression patterns in T-LBL remain poorly understood. Procedure Children with T-LBL with disseminated disease were registered and treated on COG protocol 5971. Paraffin-embedded tumor tissue was obtained at diagnosis for immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) studies. We determined the pattern and intensity of staining for c-Myc, Skp2, Mib-1, p53, TCL-1, bcl-2, and bcl-6 proteins by IHC and c-Myc, p53, bcl-2, bcl-6, and TCR ,/, molecular alterations by FISH in 22 pediatric T-LBL cases. Results The majority of T-LBL samples expressed Mib-1 (59%) and c-Myc (77%) proteins in greater than 50% of the cells, but Skp2 (14%), p53 (14%), and bcl-2 (23%) expression was less common. FISH studies demonstrated 18% gains and 10% losses in c-Myc, 16% gains in p53, 12% gains and 6% losses in bcl-2, and 6% gains and 19% losses in bcl-6 with little direct correlation between the IHC and FISH studies. Conclusions Childhood T-LBL is a highly proliferative tumor associated with enhanced expression of c-Myc protein, but without detectable c-Myc molecular alterations. FISH studies did not identify consistent etiologies of molecular dysregulation, and future studies with other molecular approaches may be required to elucidate the molecular pathogenesis of childhood T-LBL. Pediatr Blood Cancer 2008;51:489,494. © 2008 Wiley-Liss, Inc. [source] Proteome analysis of ventral midbrain in MPTP-treated normal and L1cam transgenic micePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 6 2008Madeleine Diedrich Abstract Treatment of mice by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridene hydrochloride (MPTP) is a well established animal model for Parkinson's disease (PD), while overexpression of L1 cell adhesion molecule (L1cam) has been proposed to attenuate the degeneration of dopaminergic neurons induced by MPTP. To gain insight into the role of L1cam in the pathomechanism of PD, we investigated protein expression patterns after MPTP-treatment in both C57BL/6 (wild-type) and transgenic mice overexpressing L1cam in astrocytes. Our results showed that during the acute phase, proteins in functional complexes responsible for mitochondrial, glycolysis, and cytoskeletal function were down-regulated in MPTP-treated wild-type mice. After a recovery phase, proteins that were down-regulated in the acute phase reverted to normal levels. In L1cam transgenic mice, a much higher number of proteins was altered during the acute phase and this number even increased after the recovery phase. Many proteins involved in oxidative phosphorylation were still down-regulated and glycolysis related protein were still up-regulated. This pattern indicates a lasting severely impaired energy production in L1cam mice after MPTP treatment. [source] Prevalidation of potential protein biomarkers in toxicology using iTRAQÔ reagent technologyPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 10 2007Matthias Glückmann Abstract Today, toxicoproteomics still relies mainly on 2-DE followed by MS for detection and identification of proteins, which might characterize a certain state of disease, indicate toxicity or even predict carcinogenicity. We utilized the classical 2-DE/MS approach for the evaluation of early protein biomarkers which are predictive for chemically induced hepatocarcinogenesis in rats. We were able to identify statistically significantly deregulated proteins in N -nitrosomorpholine exposed rat liver tissue. Based on literature data, biological relevance in the early molecular process of hepatocarcinogenicity could be suggested for most of these potential biomarkers. However, in order to ensure reliable results and to create the prerequisites necessary for integration in routine toxicology studies in the future, these protein expression patterns need to be prevalidated using independent technology platforms. In the current study, we evaluated the usefulness of iTRAQÔ reagent technology (Applied Biosystems, Framingham, USA), a recently introduced MS-based protein quantitation method, for verification of the 2-DE/MS biomarkers. In summary, the regulation of 26 2-DE/MS derived protein biomarkers could be verified. Proteins like HSP 90-beta, annexin A5, ketohexokinase, N -hydroxyarylamine sulfotransferase, ornithine aminotransferase, and adenosine kinase showed highly comparable fold changes using both proteomic quantitation strategies. In addition, iTRAQ analysis delivered further potential biomarkers with biological relevance to the processes of hepatocarcinogenicity: e.g. placental form of glutathione S-transferase (GST-P), carbonic anhydrase, and aflatoxin B1 aldehyde reductase. Our results show both the usefulness of iTRAQ reagent technology for biomarker prevalidation as well as for identification of further potential marker proteins, which are indicative for liver hepatocarcinogenicity. [source] The power of cooperative investigation: Summary and comparison of the HUPO Brain Proteome Project pilot study resultsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 18 2006Kai A. Reidegeld Abstract Within the pilot phase of the HUPO Brain Proteome Project, nine participating laboratories analysed human (epilepsy and/or post mortem material) and mouse brain samples (embryonic, juvenile and adult), respectively, using a variety of different state of the art techniques. Thirty-seven different analytical approaches were accomplished. Of these analyses, 17 were done differentially, i.e. the protein expression patterns of the different samples (human or mouse) were compared. A catalogue of all proteins present in the respective sample was built in 20 analyses (mapping). All data were collected in the Data Collection Center in Bochum, Germany, and were reprocessed according to thoroughly defined parameters. In this report, a summary of all results and inter-laboratory comparisons with respect to the number of identified proteins, the analysed organism, and the used techniques is presented. [source] Pseudomonas putida KT2440 responds specifically to chlorophenoxy herbicides and their initial metabolitesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 11 2006Dirk Benndorf Dr. Abstract Pseudomonas putida,KT2440 is often used as a model to investigate toxicity mechanisms and adaptation to hazardous chemicals in bacteria. The objective of this paper was to test the impact of the chlorophenoxy herbicides 2,4-dichlorophenoxyacetic acid,(2,4-D) and 2-(2,4-dichlorophenoxy)propanoic acid,(DCPP) and their metabolites 2,4-dichlorophenol,(DCP) and 3,5-dichlorocatechol,(DCC), on protein expression patterns and physiological parameters. Both approaches showed that DCC has a different mode of action and induces different responses than DCPP, 2,4-D and DCP. DCC was the most toxic compound and was active as an uncoupler of oxidative phosphorylation. It repressed the synthesis of ferric uptake regulator (Fur)-dependent proteins, e.g. fumarase,C and L -ornithine N5-oxygenase, which are involved in oxidative stress response and iron uptake. DCPP, 2,4-D and DCP were less toxic than DCC. They disturbed oxidative phosphorylation to a lesser extent by a yet unknown mechanism. Furthermore, they repressed enzymes of energy-consuming biosynthetic pathways and induced membrane transporters for organic substrates. A TolC homologue component of multidrug resistance transporters was found to be induced, which is probably involved in the removal of lipophilic compounds from membranes. [source] Proteins differentially expressed in response to nicotine in five rat brain regions: Identification using a 2-DE/MS-based proteomics approachPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 10 2006Yoon Y. Hwang Abstract To determine protein expression patterns within the central nervous system,(CNS) in response to nicotine, 2-DE/MS was performed on samples from five brain regions of rats that had received nicotine bitartrate by osmotic minipump infusion at a dose of 3.15,mg/kg/day for 7,days. After spot matching and statistical analysis, 41,spots in the amygdala, 49 in the nucleus accumbens,(NA), 46 in the prefrontal cortex (PFC), 36 in the striatum, and 28 in the ventral tegmental area,(VTA) showed significant differences in the nicotine-treated compared with control samples. Using MALDI-TOF,MS peptide fingerprinting, 14,proteins in the amygdala, 11 in the NA, 19 in the PFC, 13 in the striatum, and 19 in the VTA were identified. Several proteins (e.g. dynamin,1, laminin receptors, aldolase,A, enolase,1 alpha, Hsc70-ps1, and N -ethylmaleimide-sensitive fusion protein) were differentially expressed in multiple brain regions. By gene ontology analysis, these differentially expressed proteins were grouped into biological process categories, such as energy metabolism, synaptic function, and oxidative stress metabolism. These data, in combination with microarray analysis of mRNA transcripts, have the potential to identify the CNS gene products that show coordinated changes in expression at both the RNA and protein levels in response to nicotine. [source] Proteome characterization of human T helper 1 and 2 cellsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 1 2004Kirsi Rautajoki Abstract T helper (Th) cells can be polarized into two different main subtypes, Th1 and Th2 cells. Their activation is linked to the eradication of different pathogens and to dissimilar immunological dysfunctions, which implies differences also in their protein expression patterns. To identify these differences, CD4+ T cells were isolated from human cord blood, polarized in vitro to Th1 and Th2 and activated via CD3 and CD28. Cells were lysed, soluble proteins were separated with two-dimensional electrophoresis and differing protein spots were identified with peptide mass fingerprinting. The expression of 14 proteins differed in Th1 and Th2 cells after both 7 and 14 days of polarization. Twelve of the proteins could be identified, most of which are new in this context. Two proteins were differentially modified in the two cell types. Especially, N -terminal acetylation of cyclophilin A was stronger in Th1 than in Th2 cells. To compare the RNA and the protein levels of the identified genes, mRNA expression was measured with Affymetrix oligonucleotide microarrays (HG-U133A). The mRNA and protein expression level correlated only in six cases out of eleven, which highlights the complementary roles that proteomics and transcriptomics have in the elucidation of biological phenomena. [source] Dihydropyrimidinase related protein-2 as a biomarker for temperature and time dependent post mortem changes in the mouse brain proteomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 10 2003Bo Franzén Abstract Proteome analysis in the central nervous system area represents a large and important challenge in drug discovery. One major problem is to obtain representative and well characterized tissues of high quality for analysis. We have used brain tissues from normal mice to study the effect of post mortem time (up to 32 h) and temperature (4°C and room temperature) on protein expression patterns. A number of proteins were identified using mass spectrometry and potential markers were localized. One of the proteins identified, dihydropyrimidinase related protein-2 (DRP-2), occurs as multiple spots in two-dimensional electrophoresis gels. The ratio between the truncated form of DRP-2 (fDRP-2) and full length DRP-2 is suggested as an internal control that can be used as a biomarker of post mortem time and post mortem temperature between unrelated brain protein samples. Results of this study may be useful in future efforts to detect disease specific alterations in proteomic studies of human post mortem brain tissues. [source] Uterine Expression of Epidermal Growth Factor Family During the Course of Pregnancy in PigsREPRODUCTION IN DOMESTIC ANIMALS, Issue 5 2009Y-J Kim Contents To stably maintain pregnancy, several genes are expressed in the uterus. In particular, the endometrial expression of genes encoding growth factors appears to play a key role in maternal,foetal communication. The previous studies characterized the endometrial expression kinetics of the genes encoding epidermal growth factor (EGF), its receptor (EGFR), transforming growth factor-alpha (TGF-,), amphiregulin (Areg), heparin-binding (Hb) EGF and calbindin-D9k (CaBP-9k) in pigs during implantation. Here, we further characterized the expression patterns of these molecules during the entire porcine pregnancy. Porcine uteri were collected at pregnancy days (PD) 12, 15, 30, 60, 90 and 110 and subjected to RT-PCR. EGF and EGFR showed similar expression patterns, being highly expressed around implantation and then disappearing. TGF-, and Areg expression levels rose steadily until they peaked at PD30, after which they gradually decreased to PD12 levels. This Areg mRNA expression pattern was confirmed by real-time PCR and similar Areg protein expression patterns were observed. Immunohistochemical analysis of PD60 uteri revealed Areg in the glandular and luminal epithelial cells. Hb EGF was steadily expressed throughout the entire pregnancy, while CaBP-9k was expressed strongly on PD12, and then declined sharply on PD15 before recovering slightly for the remainder of the pregnancy. Thus, the EGF family may play a key role during implantation in pigs. In addition, CaBP-9k may help to maintain uterine quiescence during pregnancy by sequestering cytoplasmic Ca2+. [source] Abnormal Expression of TIMP-2, SOD, Vimentin and PAI Proteins in Cloned Bovine PlacentaeREPRODUCTION IN DOMESTIC ANIMALS, Issue 4 2009H-R Kim Contents Cloned mammals suffer from high rates of placental abnormality and foetal loss during pregnancy. We previously used 2-D gel electrophoresis and mass spectrometry for global proteomic analysis of cloned and normal bovine placentae to identify differential protein expression patterns. Here, we used Western blot analysis to confirm the expression levels of several pregnancy-related proteins putatively identified as being differentially expressed in somatic cell nuclear transfer (SCNT) vs normal bovine placentae. The expression levels of tissue inhibitor of metalloproteinase-2 (TIMP-2), its downstream protein, matrix metalloproteinase-2 (MMP-2), superoxide dismutase (SOD), vimentin and plasminogen activator inhibitor-1 (PAI) were analysed in the placentae of SCNT cloned Korean native cattle that died immediately after birth and in normal placentae obtained by AI. Our results revealed that TIMP-2 and SOD were up-regulated in SCNT placenta compared with normal placenta, whereas MMP-2 levels were comparable in cloned and normal placentae, and vimentin and PAI were significantly down-regulated in SCNT compared with normal placentae. Our results suggest that key proteins of placental development are abnormally expressed in SCNT cloned bovine placentae, probably resulting in abnormal placental function and clonal mortality. [source] Developmental and adult expression of semaphorin 2a in the cricket Gryllus bimaculatus,THE JOURNAL OF COMPARATIVE NEUROLOGY, Issue 1 2007Kristen R. Maynard Abstract Developmental guidance cues act to direct growth cones to their correct targets in the nervous system. Recent experiments also demonstrate that developmental cues are expressed in the adult mammalian nervous system, although their function in the brain is not yet clear. The semaphorin gene family has been implicated in the growth of dendrites and axons in a number of different species. While the expression of semaphorin and its influence on tibial pioneer neurons in the developing limb bud have been well characterized in the grasshopper, the expression of semaphorin 2a (sema2a) has not been explored in the adult insect. In this study we used polymerase chain reaction (PCR) with degenerate and gene-specific primers to clone part of the secreted form of sema2a from Gryllus bimaculatus. Using in situ hybridization and immunohistochemistry, we confirmed that sema2a mRNA and protein expression patterns in the embryonic cricket were similar to that seen in the grasshopper. We also showed that tibial neuron development in crickets was comparable to that described in grasshopper. An examination of both developing and adult cricket brains showed that sema2a mRNA and protein were expressed in the Kenyon cells in mushroom bodies, an area involved in learning and memory. Sema2a expression was most obvious near the apex of the mushroom body in a region surrounding the neurogenic tip, which produces neurons throughout the life of the cricket. We discuss the role of neurogenesis in learning and memory and the potential involvement of semaphorin in this process. J. Comp. Neurol. 503:169,181, 2007. © 2007 Wiley-Liss, Inc. [source] The Human Protein Atlas,a tool for pathology,THE JOURNAL OF PATHOLOGY, Issue 4 2008F Pontén Abstract Tissue-based diagnostics and research is incessantly evolving with the development of new molecular tools. It has long been realized that immunohistochemistry can add an important new level of information on top of morphology and that protein expression patterns in a cancer may yield crucial diagnostic and prognostic information. We have generated an immunohistochemistry-based map of protein expression profiles in normal tissues, cancer and cell lines. For each antibody, altogether 708 spots of tissues and cells are analysed and the resulting images and data are presented as freely available in the Human Protein Atlas (www.proteinatlas.org). The new version 4 of the atlas, including more than 5 million images of immunohistochemically stained tissues and cells, is based on 6122 antibodies, representing 5011 human proteins encoded by approximately 25% of the human genome. The gene-centric database includes a putative classification of proteins in various protein classes, both functional classes, such as kinases or transcription factors and project-related classes, such as candidate genes for cancer or cardiovascular diseases. For each of the internally generated antibodies, the exact antigen sequence is presented, together with a visualization of application-specific validation data, including a protein array assay, western blot analysis, immunohistochemistry and, in most cases, immunofluorescent-based confocal microscopy. The updated version also includes new search algorithms to allow complex queries regarding expression profiles, protein classes and chromosome location. Thus, the presented Human Protein Atlas provides a resource for pathology-based biomedical research, including protein science and biomarker discovery. Copyright © 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source] Proteomic analysis by two-dimensional electrophoresis to identify the normal human chondrocyte proteome stimulated by tumor necrosis factor , and interleukin-1,ARTHRITIS & RHEUMATISM, Issue 3 2010Berta Cillero-Pastor Objective To determine the intracellular proteome of normal human chondrocytes stimulated with interleukin-1, (IL-1,) and tumor necrosis factor , (TNF,) and to ascertain differences in the protein expression patterns of these 2 cytokines. Methods Normal human knee cartilage chondrocytes were incubated for 48 hours without stimulation or stimulated with IL-1, (5 ng/ml) or with TNF, (10 ng/ml). For each culture condition, protein extracts from 4 normal subjects were pooled and resolved using 2-dimensional electrophoresis. Protein spots were visualized with Sypro stain, and qualitative and quantitative analyses were performed using PDQuest software. Protein spots were then identified by mass spectrometry, using matrix-assisted laser desorption ionization,time-of-flight/time-of-flight technology. Results We identified 37 spots by mass spectrometry (MS) or by MS/MS, corresponding to 35 different proteins. In IL-1,,stimulated chondrocytes, IL-1, was found to modulate 22 proteins, as compared with unstimulated chondrocytes. All of these proteins except connective tissue growth factor (CCND2) were up-regulated. Proteins involved in cellular metabolism and energy (23%) that were up-regulated or induced by IL-1, included nicotinamide phosphoribosyltransferase, long-chain fatty acid,coenzyme A ligase 4, ,-aminolevulinic acid dehydratase, triosephosphate isomerase, and an isoform of glyceraldehyde-3-phosphate dehydrogenase. In TNF,-stimulated chondrocytes, TNF, was found to modulate 20 proteins, as compared with unstimulated chondrocytes. All of these except chitinase 3,like 1 (cartilage glycoprotein 39), proteasome activator complex subunit 2, and G3PDH, were up-regulated. Eighteen proteins were differently modulated by IL-1, and TNF,. Of these, 45% were related to metabolism. Conclusion IL-1, and TNF, induce different profiles of intracellular protein expression in healthy human chondrocytes. Most of the proteins that are differently regulated are proteins that are implicated in the generation of cellular energy and in glycolysis. [source] Insights into the relation between mRNA and protein expression patterns: I. theoretical considerationsBIOTECHNOLOGY & BIOENGINEERING, Issue 7 2003Amit Mehra Abstract Translation is a central cellular process in every organism and understanding translation from the systems (genome-wide) perspective is very important for medical and biochemical engineering applications. Moreover, recent advances in cell-wide monitoring tools for both mRNA and protein levels have necessitated the development of such a model to identify parameters and conditions that influence the mapping between mRNA and protein expression. Experimental studies show a lack of correspondence between mRNA and protein expression profiles. In this study, we describe a mechanistic genome-wide model for translation that provides mapping between changes in mRNA levels and changes in protein levels. We use our model to study the system in detail and identify the key parameters that affect this mapping. Our results show that the correlation between mRNA and protein levels is a function of both the kinetic parameters and concentration of ribosomes at the reference state. In particular, changes in concentration of free and total ribosomes in response to a perturbation; changes in initiation and elongation kinetics due to competition for aminoacyl tRNAs; changes in termination kinetics; average changes in mRNA levels in response to the perturbation; and changes in protein stability are all important determinants of the mapping between mRNA and protein expression. © 2003 Wiley Periodicals, Inc. [source] Unique Molecular Characteristics of Pediatric Myxopapillary EpendymomaBRAIN PATHOLOGY, Issue 3 2010Valerie N. Barton Abstract Myxopapillary ependymoma (MEPN) generally can be cured by gross total surgical resection and usually manifest a favorable prognosis. However, surgery is less curative in tumors that are large, multifocal or extend outside the thecal sac. Late recurrences may occur, particularly in pediatric patients. The role of adjuvant therapy is unclear in the clinical management of recurrent tumors. Clinical trial design requires a better understanding of tumor biology. Unique molecular features of MEPN were investigated by using microarray technology to compare the gene expression of five pediatric MEPN to 24 pediatric intracranial ependymoma (EPN). The upregulation of three genes of interest, homeobox B13 (HOXB13), neurofilament, light polypeptide (NEFL) and PDGFR,, was further studied by immunohistochemistry in a larger cohort that included adult MEPN and EPN specimens. Protein expression in MEPN was compared to subependymoma, spinal EPN, intracranial EPN and normal fetal and adult ependyma. Immunoreactivity for HOXB13, NEFL and PDGFR, was strongest in MEPN and virtually absent in subependymoma. Spinal and intracranial EPN generally expressed weak or focal staining. MEPN manifests unique gene and protein expression patterns compared to other EPNs. Aberrant expression of HOXB13 suggests possible recapitulation of developmental pathways in MEPN tumorigenesis. PDGFR, may be a potential therapeutic target in recurrent MEPN. [source] Cell volume and rate of proliferation, but not protein expression pattern, distinguish pup/intimal smooth muscle cells from subcultured adult smooth muscle cellsCELL PROLIFERATION, Issue 5 2001E. McKilligin Smooth muscle cells from neonatal rats and from injured blood vessels grow with a characteristic cobblestone morphology that distinguishes them from adult smooth muscle cells. This has led to the proposition that there are two distinct types of smooth muscle cells with different proliferative capacity. Here we systematically compare the properties of subcultured adult smooth muscle cells in culture and clonal lines of cobblestone smooth muscle cells from both neonatal rats and injured vessels. The cobblestone smooth muscle cells have a significantly smaller average cell volume, estimated using two different flow cytometry measurements. However, the two types of smooth muscle cells have indistinguishable protein expression patterns when the levels of more than 20 different proteins (including cytoskeletal proteins, matrix proteins, cytokines, cytokine receptors, adhesion molecules and enzymes) are measured by quantitative immunofluorescence. Furthermore, in contrast to previous observations, we demonstrate that both types of smooth muscle cells secrete a powerful mitogenic activity. The higher cell density achieved by the cobblestone smooth muscle cells in culture was responsible for the earlier reports that this mitogenic activity was secreted only by cobblestone smooth muscle cells. We conclude that many of the differences seen between cobblestone smooth muscle cells and adult smooth muscle cells in vitro (proliferation rate, morphology, protein expression pattern, secretion of mitogenic activity) could be attributable to a stable difference in the median cell volume of the cultures. [source] | ||||||||||||||||