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Results for “differential analysis”

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At least 595 records · Page 33Linked to original sources
PubMed2003

Proteomic analysis of differential protein expression induced by ultraviolet light radiation in HeLa cells.

Cells treated with ultraviolet (UV) radiation undergo cell cycle arrest at the S-phase and G1/S boundary, allowing DNA repair to occur. Several proteins such as replication protein A and DNA-dependent protein kinase have been suggested to be involved in UV-induced inhibition of DNA replication. However, the role of these proteins in inhibiting DNA replication remains unknown. Other proteins may play important roles in modulating functions of these proteins in response to UV-irradiation. To understand the broad range of proteins involved in this inhibition, we carried out a systematic study to identify specific proteins involved in UV-induced replication arrest using two-dimensional gel electrophoresis and mass spectrometry. Unique changes in protein expression level for 31 proteins were observed over a 24-hour time course, including calgizzarin, cyclophilin A, and macrophage migration inhibitory factor. The expression level changes of these proteins are dynamically correlated to DNA replication activity, suggesting involvement of these proteins in modulating DNA replication and repair activities. This proteomic approach provides opportunities to gain insights into the mechanism by which DNA replication is inhibited.

DNA Repair↗
PubMed2004

Differential proteomic analysis of Aeromonas salmonicida outer membrane proteins in response to low iron and in vivo growth conditions.

Aeromonas salmonicida subsp. salmonicida is the etiological agent of furunculosis, a serious infectious disease of salmonids. Bacterial phenotypes are known to change in vivo compared to the in vitro state. Proteomic analysis of in vivo phenotypes is usually not possible due to insufficient biomass. Using an in vivo growth chamber model, the pathogenic fish bacterium A. salmonicida was cultured in pure culture in vivo in its host, the Atlantic salmon, to obtain sufficient biomass to allow proteomic analysis. Growth of A. salmonicida under in vitro iron-restricted conditions resulted in the expression of outer membrane proteins of 73, 76 and 85 kDa, which were not present when grown under in vitro iron-replete conditions. Mass spectrometry analysis identified the 73 kDa protein as a colicin receptor, the 76 kDa protein as an outer membrane heme receptor, and the 85 kDa protein as a ferric siderophore receptor. When cultured in vivo, A. salmonicida up-regulated the identical 73, 76 and 85 kDa proteins. The results of this study also suggest, at least with respect to the outer membrane proteins, that the in vitro iron-restricted growth model largely reproduces the results obtained from growth of A. salmonicida within the peritoneal cavity of salmon.

Aeromonas↗
PubMed2004

Proteomic analysis of differential protein expression in neuropathic pain and electroacupuncture treatment models.

Acupuncture has long been used for pain relief. Although recent studies have shown that acupuncture can reduce neuropathic pain, the mechanism of this effect is not clear and little information is available regarding proteins that are involved in the development of neuropathic pain and the effects of acupuncture. We have developed an animal model for neuropathic pain using young adult male Sprague-Dawley rats. The model was confirmed by behavioral tests. Electroacupuncture (EA) treatment was applied to Zusanli (ST36) of neuropathic pain model to examine the analgesic effect of EA. The protein expression profile of the hypothalamus in both neuropathic pain and EA treatment models was analyzed using two-dimensional electrophoresis-based proteomics. We detected thirty-six proteins that were differentially expressed in the neuropathic pain model compared with normal rats and that restored to normal expression levels after EA treatment. Twenty-one of these proteins were identified in the MS-FiT database and are involved in a number of biological processes, including inflammation, enzyme metabolism and signal transduction. Potential applications of our results include the identification and characterization of signaling pathways involved in EA treatment and further exploration of the role of selected identified proteins in the animal model.

Animals↗
PubMed2004

Differential proteome analysis and mass spectrometric characterization of germ line development-related proteins of Caenorhabditis elegans.

Proteome maps and differences of protein patterns of the synchronized larval stage L4 of the temperature-sensitive Caenorhabditis elegans (C. elegans) glp-1 mutant (e2144ts) were investigated after cultivation at 15 degrees C (developing a normal phenotype) or 25 degrees C (developing a mutated phenotype) by two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization (MALDI)-mass spectrometry. From the 183 identified protein spots six proteins were found differently expressed. The Vit-6 vitellogenin (CE28594), the hypothetical 17.2 protein (CE25224), the hypothetical 17.4 protein (CE16999), and the heat shock protein 16 kDa (CE14249) were more abundant when growing worm cultures at 25 degrees C. By contrast, the nucleoside diphosphate kinase (CE09650) was found increased at 15 degrees C. Most notably, the eukariotic initiation factor 5A-1 (CE00503), highly abundant at 15 degrees C, was not present in cultures grown at 25 degrees C. Its absence at 25 degrees C can not be attributed to lack of the enzymatic machinery that is necessary for hypusinylation. Instead, a direct downstream effect of the lack of functionality of GLP-1 may cause the expression of this protein. The yolk proteins 115 kDa and 88 kDa were attributed by mass spectrometric protein structure analysis as C-terminal and N-terminal fragments of the Vit-6 vitellogin protein (CE28594), respectively. The cleavage site between both derivatives was located between R764 and A768. A conflict in the database sequences at amino acid positions 1622 and 1623 of vitellogenin-6 was solved by mass spectrometric sequence analysis. The combination of 2-DE with mass spectrometry enabled the identification of mutation-associated differences on somatic gonadal cell and germ line cell development-associated proteins.

Amino Acid Sequence↗
PubMed2004

Proteome analysis of differential protein expression in allergen-induced asthmatic mice lung after dexamethasone treatment.

Asthma has become substantially more prevalent in recent decades and is one of the foremost contributors to morbidity and mortality in industrialized countries. Corticosteroids are among the most effective medications for the treatment of asthma, but some patients do not respond well to corticosteroid treatment. In this study, we characterized the responses to an allergen and identified potential molecular targets of dexamethasone (Dex) treatment in acute asthma. Female BALB/c mice sensitized to ovalbumin (OVA) were challenged with aerosolized OVA for 1 week. During the challenge period, mice were treated daily with Dex by intraperitoneal injection. Phosphate-buffered saline treated and non-challenged mice served as control. Histological evaluation of OVA-induced mice revealed airway inflammation and goblet cell hyperplasia. In addition, interleukin 4 levels and interferon-gamma levels were increased and decreased, respectively. These changes were moderated by Dex treatment. Protein expression profiles were compared in each experimental group by two-dimensional gel electrophoresis and identified using matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry. Some proteins were increased, while others were decreased by Dex treatment. These results indicated that the regulation of protein expression might play a role in the immunological and pathological development of asthma and could be targeted for therapeutic intervention. These results may assist in the development of quantitative diagnostic markers to monitor disease progression or responses to therapy using proteomic approaches.

Animals↗
PubMed2005

Proteomic analysis of differentially expressed proteins between the male and female worm of Schistosoma japonicum after pairing.

Identification of differentially expressed proteins between the male and female worm of Schistosoma japonicum may provide new insights into the development of schistosomes, especially the molecular mechanism of female worm maturation induced by the male worm after pairing. Comparative two-dimensional gel electrophoresis (2-DE) and mass spectrometry were employed to separate and identify differentially expressed proteins between the male and female worm after pairing. Soluble and hydrophobic proteins from egg, schistosomulum (14 days), and female and male worms at adult stage (42 days) were separated by a sequential extraction method followed by 2-DE and 2-DE images were constructed. There were 1016 +/- 67, 1808 +/- 89, 1142 +/- 45 and 1288 +/- 32 spots detected for soluble proteins and 1425 +/- 108, 952 +/- 59, 847 +/- 75 and 965 +/- 69 spots for hydrophobic proteins from egg, schistosomulum, and adult stage female and male worms, respectively. The differentially and uniquely expressed proteins from male and female worms after pairing (42 days) include 41 +/- 4 and 23 +/- 2 unique spots for soluble proteins, and 11 +/- 1 and 26 +/- 3 unique spots for hydrophobic proteins, respectively. Matrix-assisted laser desorption/ionization-time of flight and electrospray ionization-tandem mass spectrometry were employed to analyze 12 unique spots from the female worm and 16 unique spots from the male worm for peptide mass fingerprinting and sequencing. The results showed that the main functions of these differentially expressed proteins were in signal transduction, metabolism and transcriptional regulation etc. Comparison of the schistosomes proteome between male and female worms may permit the identification of protein candidates for the development of vaccines or new targets for drug development against schistosomiasis.

Animals↗
PubMed2005

Differential proteomic analysis of proteins in wheat spikes induced by Fusarium graminearum.

Scab, caused by Fusarium graminearum, is a serious spike disease in wheat. To identify proteins in resistant wheat cultivar Wangshuibai induced by F. graminearum infection, proteins extracted from spikes 6, 12 and 24 h after inoculation were separated by 2-DE. Thirty protein spots showing 3-fold change in abundance when compared with treatment without inoculation were characterized by MALDI-TOF MS and matched to proteins by querying the mass spectra in protein databases or the Triticeae EST translation database. Based on their volume profiles, these proteins were classified into four categories. The first one fell off rapidly at the initial inoculation and then rose at 12 or 24 hai, the second one decreased considerably after inoculation and remained at low level, the third one rose at the initial inoculation and then declined at 12 or 24 hai, the forth one showed steady increase after inoculation and maintained at a high level. Many of the proteins identified in the first two categories are related to carbon metabolism and photosynthesis. While most of proteins identified in the last two categories are related to stress defense of plants, indicating that proteins associated with the defense reactions were activated or translated shortly after inoculation.

Electrophoresis, Gel, Two-Dimensional↗
PubMed2006

The tegument surface membranes of the human blood parasite Schistosoma mansoni: a proteomic analysis after differential extraction.

The blood fluke Schistosoma mansoni can live for years in the hepatic portal system of its human host and so must possess very effective mechanisms of immune evasion. The key to understanding how these operate lies in defining the molecular organisation of the exposed parasite surface. The adult worm is covered by a syncytial tegument, bounded externally by a plasma membrane and overlain by a laminate secretion, the membranocalyx. In order to determine the protein composition of this surface, the membranes were detached using a freeze/thaw technique and enriched by sucrose density gradient centrifugation. The resulting preparation was sequentially extracted with three reagents of increasing solubilising power. The extracts were separated by 2-DE and their protein constituents were identified by MS/MS, yielding predominantly cytosolic, cytoskeletal and membrane-associated proteins, respectively. After extraction, the final pellet containing membrane-spanning proteins was processed by liquid chromatographic techniques before MS. Transporters for sugars, amino acids, ions and other solutes were found together with membrane enzymes and proteins concerned with membrane structure. The proteins identified were categorised by their function and putative location on the basis of their homology with annotated proteins in other organisms.

Animals↗
PubMed2007

A comparative proteome and phosphoproteome analysis of differentially regulated proteins during fertilization in the self-incompatible species Solanum chacoense Bitt.

We have used 2-DE for a time-course study of the changes in protein and phosphoprotein expression that occur immediately after fertilization in Solanum chacoense. The phosphorylation status of the detected proteins was determined with three methods: in vivo labeling, immunodetection, and phosphoprotein-specific staining. Using a pI range of 4-7, 262 phosphorylated proteins could be mapped to the 619 proteins detected by Sypro Ruby staining, representing 42% of the total proteins. Among these phosphoproteins, antibodies detected 184 proteins from which 78 were also detected with either of the other two methods (42%). Pro-Q Diamond phosphoprotein stain detected 111 proteins, of which 76 were also detected with either of the other two methods (68%). The 32P in vivo labeling method detected 90 spots from which 78 were also detected with either of other two methods (87%). On comparing before and after fertilization profiles, 38 proteins and phosphoproteins presented a reproducible change in their accumulation profiles. Among these, 24 spots were selected and analyzed by LC-MS/MS using a hybrid quadrupole-TOF (Q-TOF) instrument. Peptide data were searched against publicly available protein and EST databases, and the putative roles of the identified proteins in early fertilization events are discussed.

Fertilization↗
PubMed2001

Genomic analysis of differentially expressed genes in liver and biliary epithelial cells of patients with primary biliary cirrhosis.

The characterization of differentially expressed genes provides a powerful tool for identifying molecules that may be involved in the pathogenesis of disease. We have used two independent techniques to identify overexpressed transcripts in bile duct cells and in liver from patients with primary biliary cirrhosis (PBC). In the first method, we used suppressive subtractive hybridization to compare mRNA from isolated PBC bile duct epithelial cells (BECs) to normal BECs and identified 71 clones as transcribed at higher levels in PBC-BECs. Amongst these clones, 62/71 had matches in a non-redundant nucleotide database and 9/71 had matches in an EST database. Of the 62 clones, 51/62 include a complexity of genes involved in cell proliferation, signal transduction, transcription regulation, RNA processing, carbohydrate metabolism and hypothetical/unknown proteins; 4/62 were identified as interstitial collagenase and collagenase precursors, 4/62 as ribosomal proteins, 3/62 as mitochondrial DNA. The mitochondrial cDNA sequences included cytochrome c oxidase, Wnt-13, and the pHL gene, a c-myc oncogene containing coxIII sequence. In the second method, we constructed cDNA libraries from three different PBC livers and sequenced a total of 12,324 independent clones. These 12,324 clones underwent virtual subtraction with 2,814,148 independent clones from Incyte LifeSeq libraries. Twenty one sequences were identified as unique to PBC liver. Collectively, these approaches identified a number of genes involved in signalling, RNA processing, mitochondrial function, inflammation, and fibrosis. Interestingly, both Wnt-13 and Notch transcripts are overexpressed in PBC liver. Further studies are needed to focus on the significance of these genes during the natural history of disease.

Bile Ducts↗
PubMed1999

Analysis of differential effects of Pb2+ on protein kinase C isozymes.

Protein kinase C has been implicated as a cellular target for Pb2+ toxicity. We have previously proposed that Pb2+ modulates PKC activity by interacting with multiple sites within the enzyme. In order to further characterize the Pb-PKC interactions we compared the effects of Pb2+ on the CA-dependent and -independent protein kinase C isozymes using recombinant human PKC-alpha, PKC-epsilon, and PKC-zeta as well as the catalytic fragment of bovine brain protein kinase C, the PKC-M. The results demonstrate that, whereas at pM concentrations Pb2+ activates PKC-alpha half maximally (KAct approximately 2 pM), it has no effect on PKC-epsilon, PKC-zeta, or PKC-M activities. The activation of PKC-alpha by Pb2+ is additive with Ca2+ in a manner indicating interaction with half of the calcium activation sites. In the micromolar range of concentrations, Pb2+ inhibits all PKCs with estimated K0.5 of 1.0, 2.3, 28, and 93 microM for PKC-M, PKC-alpha, PKC-epsilon, and PKC-zeta, respectively. Examination of Pb2+ effects on PKC-M kinetics indicates a mixed type inhibition with respect to ATP and noncompetitive inhibition with respect to histone. Taken together with the results of our previous study (Tomsig and Suszkiw, J. Neurochem. 64, 2667-2673, 1995) and the evidence for the existence of two Ca2+ coordination sites Ca1 and Ca2 within the C2 domain (Shao et al., Science [Washington, D.C.] 273, 248-251, 1996), the results of the current study provide further support for a multisite Pb-PKC interaction scheme wherein lead (1) partially activates the enzyme through pM-affinity interactions with the Ca1 site and inhibits the divalent cation-dependent activity through nM-affinity interactions with Ca2 site in the C2 domain and (2) inhibits the constitutive kinase activity through microM-affinity interactions with the catalytic domain. The concentration dependence of the differential effects of Pb2+ on the calcium-dependent and -independent PKCs underscores the importance of the C2 motif as a high affinity molecular target for Pb2+.

Amino Acid Sequence↗
PubMed2001

Comparative expressed-sequence-tag analysis of differential gene expression profiles in BmNPV-infected BmN cells.

To compare the gene expression profiles of uninfected and Bombyx mori nucleopolyhedrovirus (BmNPV)-infected BmN cells, we constructed four cDNA libraries for mock-infected cells, and cells at 2, 6, and 12 h postinfection (h.p.i.). A total of 2645 partial sequences obtained for expressed-sequence-tags (ESTs) from the libraries were categorized using BLAST searches of the public database and the BmNPV genome sequence. The following proportions of BmNPV-derived ESTs were observed: 0.4, 4.5, and 57% at 2, 6, and 12 h.p.i, respectively. Moreover, 31 BmNPV open reading frames (ORFs) were newly identified for transcripts and the baculovirus-repeated ORFs (bro) showed the highest levels of expression in the 12 h.p.i. library. Most of the host genes decreased in number as the infection progressed. However, several, including cytochrome c oxidase 1, increased in the late stages of infection. Two apoptosis-related host genes were also identified.

Amino Acid Sequence↗
PubMedDate not supplied

In vitro modulation of the metastatic phenotype. I. Analysis of differentiation forms of the B16 melanoma expressing Met-72 determinants and metastatic activity.

In vitro cultures of a highly metastatic B16 melanoma clone (BL6-10) were found to undergo dramatic changes in morphology and differentiation upon transfer to another culture medium. Specifically, BL6-10 melanoma cells which had been originally selected and adapted for growth in Eagles' Hanks' amino acid supplemented media with 10 per cent newborn calf serum were amelanotic and epitheliod in shape. When these cells were shifted into Dulbecco's modified Eagles medium with 10 per cent fetal calf serum, they became highly melanotic and of spindle/dendritic morphology within 4 days of culture. These morphological changes as well as other parameters were all characteristic of established criteria of melanoma differentiation. Alterations in the differentiation state of our highly metastatic variant, BL6-10, did not result in any change in tumorigenicity but did have profound effects on metastatic potential. All of the morphological and functional characteristics of the differentiated melanoma were found to be reversible by re-plating the cells in their original growth medium and 4 days of in vitro growth. These studies have allowed us to follow and more firmly establish Met-72 antigen expression as a surface marker for metastatic cells of the B16 melanoma, and have provided direct experimental evidence that the less differentiated, Met-72 positive melanoma form is the dominant cell type capable of metastatic potential.

Animals↗
PubMed1981

High resolution analysis and differential condensation in RBA-banded human chromosomes.

Human prophase, premetaphase, and mid-metaphase chromosomes are prepared and analyzed using the thymidine cell synchronization technique and R-banding patterns (RBA). Haploid sets with 700-1000 bands can be demonstrated. Sequences of chromosomes of different degrees of condensation are helpful for a better understanding and classification of regions of extended chromosomes. A considerable variation in the condensation of parts of homologous chromosomes is reflected in the variability of the arm ratio. This differential condensation of chromosomes is entirely effected by variation of the degree of condensation in AT rich interbands and can be attributed to the degree of labeling by BrdU.

Acridine Orange↗
PubMed1982

On the development of the spinal cord of the clawed frog, Xenopus laevis. II. Experimental analysis of differentiation and migration.

In order to determine the time and site of origin and the final location of various cell groups in the spinal cord, tadpoles of Xenopus laevis, ranging from stage 48 to stage 56 were treated with tritiated thymidine and sacrificed at various stages from 49 to 66 (stages according to Nieuwkoop and Faber (1967). From the poorly developed matrix at stage 48-49 not only ventral horn cells, but also neuroblasts of the intermediate zone and the dorsal horn arise. Both the matrix and the ventricle expand in a dorsal direction. From the well-developed matrix at stage 54, in which the mitotic activity is almost exclusively confined to its dorsal part, mainly cells of the dorsal horn develop. However, this later-stage matrix also gives rise to a considerable number of neuroblasts, which become located in the central parts of the intermediate zone and the ventral horn. Generally the later-born cells come to lie dorsomedially to the older ones. The neuroblasts of the lateral motor column, however, migrate through and settle ventrolaterally to their predecessors. Our observations do not support the basal plate-alar plate concept of His (1893).

Animals↗