PANTHER Database Research Articles

PANTHER-PSEP: predicting disease-causing genetic variants using position-specific evolutionary preservation.

PANTHER-PSEP is a new software tool for predicting non-synonymous genetic variants that may play a causal role in human disease. Several previous variant pathogenicity prediction methods have been proposed that quantify evolutionary conservation among homologous proteins from different organisms. PANTHER-PSEP employs a related but distinct metric based on 'evolutionary preservation': homologous proteins are used to reconstruct the likely sequences of ancestral proteins at nodes in a phylogenetic tree, and the history of each amino acid can be traced back in time from its current state to estimate how long that state has been preserved in its ancestors. Here, we describe the PSEP tool, and assess its performance on standard benchmarks for distinguishing disease-associated from neutral variation in humans. On these benchmarks, PSEP outperforms not only previous tools that utilize evolutionary conservation, but also several highly used tools that include multiple other sources of information as well. For predicting pathogenic human variants, the trace back of course starts with a human 'reference' protein sequence, but the PSEP tool can also be applied to predicting deleterious or pathogenic variants in reference proteins from any of the ∼100 other species in the PANTHER database. PANTHER-PSEP is freely available on the web at http://pantherdb.org/tools/csnpScoreForm.jsp Users can also download the command-line based tool at ftp://ftp.pantherdb.org/cSNP_analysis/PSEP/ CONTACT: pdthomas@usc.edu Supplementary data are available at Bioinformatics online.

Hindlimb Muscles of 17.5–18.5 dpc Mice Double Null for MyoD and Trp53 Appear Indistinguishable from Muscles of Mice Null for Either Gene

It has been shown by others that mice with either a MyoD null (MN) or Trp53 null (PN) genotype undergo myogenesis resulting in perinatal hindlimb muscle essentially indistinguishable from that of wild type (WT), while myogenic cells of the same null genotypes undergo impaired differentiation in culture. Both MyoD and Trp53 act through p21 and pRb to influence cell cycle withdrawal and differentiation. We hypothesized that redundancies in the activities of Trp53 and MyoD might rescue in vivo myogenesis in animals null for either protein, in which case animals double null (DN) for MyoD and Trp53 should demonstrate impaired hindlimb myogenesis. We bred PN and MN animals and their progeny to obtain WT, MN, PN, and DN pups. At 17.5–18.5 dpc, all possible genotypes, including DN, and both genders were present with no significant difference from the expected Mendelian distributions. Survival to adulthood was significantly reduced among MN and DN animals (P<0.001), with reduction in DN more severe. The effect of PN on survival was unclear. Location, shapes, and domains of myonuclei and alignment, continuity, branching pattern, and number and diameter of the muscle fibers were compared between MN and DN siblings, using paraffin‐embedded, hematoxylin and eosin‐stained sections of extensor digitorum longus (EDL) fast‐twitch and soleus (SOL) slow‐twitch muscles. DN muscles had no detectable deficiencies compared to the muscles of MN (N = 7 sibling sets of each muscle type). mRNA levels were compared across all 4 genotypes using NimbleGen Systems 60‐mer expression arrays, 2 arrays per genotype. Two biologically independent cDNA samples were prepared per genotype, each from pools of RNA isolated from hindlimb muscle of 5–6 mice. cDNA labeling, array hybridization, and initial normalization of data were carried out by Nimblegen. Subsequent analysis using ArrayStar software required substantial accommodation for scatter. 4417 calls (~2873 genes) with >2‐fold difference in expression level between the two arrays of any one genotype were removed from analysis and the remaining 38,169 calls (~24,831 genes) were rescaled by quantile ranking. Of genes eliminated, ~135 were involved in muscle function (Panther Database, PD), but only 2 of the 135 were eliminated because of scatter unique to the DN arrays. Thus increased scatter of muscle gene expression cannot be attributed specifically to the DN genotype. Four genes used for normalization of mRNA studies in muscle, Hprt1, Gapdh, Sf3a1, and Hadha, showed consistent levels of expression across all 4 genotypes. mRNAs from the different members of the myosin heavy chain gene family showed the expected diversity in levels of expression. Only 7 genes passed our multi‐step screening for differential expression with a >2.2 fold difference in expression between any 2 genotypes, including VIP neuropeptide hormone, and Myosin heavy chain 4 (Myh4), both involved in muscle contraction. Myh4 mRNA was elevated in MN and DN mice. Vip mRNA was elevated in the DN relative to PN but less relative to MN and WT. Thus, we found no convincing support for a model in which p53 and MyoD act as back‐ups for each other during prenatal myogenesis. It would be of interest to determine the effect of the DN on muscle regeneration and neuromuscular development, which have been shown to be abnormal in the MN mouse. Animal studies were performed in accordance with Cal State LA IACUC 06‐5.Support or Funding InformationCal State LA MBRS‐RISE 1 R25 GM61331 to C. Gutierrez; NIH‐AREA Grant #R15 AR51322‐01 to Sandra B. Sharp

Quantitative Proteomic Analysis of Rat Condylar Chondrocytes during Postnatal Development.

To investigate differentially expressed proteins in rat mandibular condylar cartilage (MCC) chondrocytes caused by initial mastication for short postnatal periods. Four groups of protein samples were extracted from primary cultured rat MCC chondrocytes, harvested from eigthy postnatal SD rats aged 1,7,14 and 28 days, with twenty in each group. Total proteins were labelled with isobaric tags for relative and absolute quantification (iTRAQ) reagents. Two-dimensional nano-high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption ionization-time-of-flight/ time-of-flight (MALDI-TOF/TOF) mass spectrometry analysis with iTRAQ technique were performed. All data were analysed by MASCOT software with the SWISSPROT protein database. Furthermore, bioinformatics and statistical analysis were performed to classify their cellular components, biological processes, molecular functions and metabolic pathway by the PANTHER database. In total, 137 differentially expressed proteins were identified during MCC growth and were assigned to one or more cellular components. According to the PANTHER analysis, a significant proportion of proteins are involved in the metabolic process, cellular process, biological regulation, developmental process and response to stimulus. The most extensive molecular function was 43% in catalytic activity. In addition, it was found that proteins in MCC chondrocytes change markedly on the growth stage of eruption of the teeth. This study provides an integrated perspective of molecular mechanisms regulating early normal postnatal growth and development of rat MCC at the protein level.

Genomic population structure and prevalence of copy number variations in South African Nguni cattle.

BackgroundCopy number variations (CNVs) are modifications in DNA structure comprising of deletions, duplications, insertions and complex multi-site variants. Although CNVs are proven to be involved in a variety of phenotypic discrepancies, the full extent and consequence of CNVs is yet to be understood. To date, no such genomic characterization has been performed in indigenous South African Nguni cattle. Nguni cattle are recognized for their ability to sustain harsh environmental conditions while exhibiting enhanced resistance to disease and parasites and are thought to comprise of up to nine different ecotypes.MethodsIllumina BovineSNP50 Beadchip data was utilized to investigate genomic population structure and the prevalence of CNVs in 492 South African Nguni cattle. PLINK, ADMIXTURE, R, gPLINK and Haploview software was utilized for quality control, population structure and haplotype block determination. PennCNV hidden Markov model identified CNVs and genes contained within and 10 Mb downstream from reported CNVs. PANTHER and Ensembl databases were subsequently utilized for gene annotation analyses.ResultsPopulation structure analyses on Nguni cattle revealed 5 sub-populations with a possible sub-structure evident at K equal to 8. Four hundred and thirty three CNVs that formed 334 CNVRs ranging from 30 kb to 1 Mb in size are reported. Only 231 of the 492 animals demonstrated CNVRs. Two hundred and eighty nine genes were observed within CNVRs identified. Of these 149, 28, 44, 2 and 14 genes were unique to sub-populations A, B, C, D and E respectively. Gene ontology analyses demonstrated a number of pathways to be represented by respective genes, including immune response, response to abiotic stress and biological regulation processess.ConclusionsCNVs may explain part of the phenotypic diversity and the enhanced adaptation evident in Nguni cattle. Genes involved in a number of cellular components, biological processes and molecular functions are reported within CNVRs identified. The significance of such CNVRs and the possible effect thereof needs to be ascertained and may hold interesting insight into the functional and adaptive consequence of CNVs in cattle.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2122-z) contains supplementary material, which is available to authorized users.

Differential proteome profiling of Helicobacter pylori-associated gastric cancer

Objective To screen for the differential expressed proteins in Helicobacter pylori-associated gastric cancer by using isobaric tags for relative and absolute quantitation(iTRAQ)labeling and liquid chromatography-electrospray ionization-tandem mass spectrometry(LC-ESI-MS/MS), and to explore the potential targets for biomarkers and biological therapy. Methods Collection the proteins of gastric epithelial cells HGC-27 and HGC-27 co-cultured with Hp, then strong cation exchange(SCX) separation after iTRAQ reagents labeled, then identification and quantitative via tandem mass spectrometry (MS/MS).The literature about differential expressed protein in Hp-associated gastric cancer by any of proteomic techniques published between January 1995 and July 2014 were selected by searching Pub Med database.The screening results of iTRAQ labeling and the reported data were meta-analyzed by PANTHER database. Results Forty-seven differential expressed proteins were identified by using iTRAQ labeling. Among the 47 proteins,37 were up-regulated and 10 were down-regulated.Seven pieces of eligible literature were obtained by information retrieval, 21 were reported by two or more labs.The iTRAQ data and the reported data meta-analyzed to get 65 differential expressed proteins.The gene ontology(GO)analysis demonstrated the major molecular functions including nucleic acid and DNA binding, catalytic activity and transferase activity as well as the cellular component organization and metabolic process in biological processes.PANTHER Protein Class, mainly for chaperone, binding protein,cytoskeletal protein, transferase and transcription factor.The differential proteins were mainly involved in Parkinson disease pathway, apoptosis signaling pathway and gonadotropin releasing hormone receptor pathway. Conclusion Screening for a variety of differential expressed proteins in Hp-associated gastric cancer may provide the potential gastric cancer-related biomarkers and therapeutic targets in early diagnosis and treatment. Key words: Stomach Neoplasms; Helicobacter pylori; Proteomics; Isobaric tags for relative and absolute quantitation

Mutations in the GnRH signaling pathway are risk factors for endometriosis

Some but not all patients with endometriosis have abnormal functioning of their GnRH axis and some but not all endometriosis patients respond to GnRH agonist therapies. The clinical variation observed may be caused by instrinsic genetic variation in these pathways. We tested 177 genes involved in human GnRH signaling for genetic association with endometriosis. Candidate genes involved in human GnRH signaling pathways were obtained using PANTHER database. 1537 Caucasian endometriosis patients were genotyped for 570 exome variants in 179 candidate genes. The frequency of heterozygous affected women was compared with published population data (n>50,000 population controls). Human Exome Beadchips (Illumina, San Diego, CA) were used for genotyping. Single marker association was tested using Fisher’s Exact Test. In silico prediction of protein function was evaluated using the Polyphen 2 database. We tested 570 exome variants representing 177 genes involved in GnRh signaling pathway that showed a causative/deleterious effect. After adjusting for multiple testing (p<8.7E-05), we discovered 5 exome variants representing 5 different genes as shown in Table 1. The top 2 significantly associated variants (CRTC1 and ARHGAP32) are predicted to be damaging to the encoded protein.Table 1CHRPositionVar AlleleEndm FreqPopulation FreqGeneFisher pOR1918876309G0.160990.08759CRTC19.51E-372.0011128839405A0.042070.02420ARHGAP328.09E-091.771877170479A0.003900.00037NFATC12.62E-0810.73112161530A0.001950.000048IGF22.37E-0740.37636075326A0.001300.000029MAPK142.28E-0544.83 Open table in a new tab Gene variants affecting the human GnRH signaling pathway are significant risk factors for predisposition to endometriosis. These gene variants are likely to contribute to the clinical variation observed in women with endometriosis.

Rare mutations in WNT signaling pathways are risk factors for endometriosis

Wnt proto-oncogenes are believed to play a role in endometriosis. Recently, genetic association studies have shown that common variants near Wnt4 are associated with endometriosis across different ethnicities. In order to test the hypothesis that variants in other genes involved in Wnt signaling pathways contribute to the pathogenesis of endometriosis, we tested exome variants found in 230 candidate genes involved in Wnt signaling for genetic association with endometriosis. A list of candidate genes involved in Wnt signaling were obtained using PANTHER database. 1537 Caucasian endometriosis patients were genotyped for 1138 rare exome variants in candidate genes. The number of heterozygous subjects observed was compared with published population data (n>50,000). 1537 patients with surgically confirmed endometriosis were tested using the Infinium HumanExome BeadChip (Illumina, San Diego, CA). Single marker association was tested using Fisher’s exact Test. In silico prediction of protein function was estimated using polyphen 2 database. We tested a total of 1138 variants representing 230 candidate Wnt genes. 88 showed causative/deleterious effect for endometriosis. After adjusting for multiple testing (p< 4.4E-05), we discovered 10 strongly associated variants in 10 distinct genes. None of these associated variants were predicted to be protein altering. The average OR among these associated variants is 10.7. Wnt signaling proteins, particularly the protocadherins, may play a major role in the pathogenesis of endometriosis.Table 1CHRPositionVar AlleleEndm FreqPopulation FreqGeneFisher pOR5140238124A0.06560.0131PCDHA107.54E-705.275140795212A0.00850.0005PCDHGA104.84E-2015.615140230533A0.00920.0020PCDHA91.94E-104.732246929692A0.06440.0399CELSR13.75E-101.651877170479A0.00390.0004NFATC12.62E-0810.735140250471A0.00200.0000PCDHA112.37E-0740.371710212619G0.00330.0004MYH131.83E-068.135140802374A0.00360.0006PCDHGA118.45E-065.911540581543G0.00490.0012PLCB21.74E-053.971668867265G0.00100.0000CDH12.41E-05Inf Open table in a new tab

Chronic Periodontitis Genome-wide Association Studies

Recent genome-wide association studies (GWAS) of chronic periodontitis (CP) offer rich data sources for the investigation of candidate genes, functional elements, and pathways. We used GWAS data of CP (n = 4,504) and periodontal pathogen colonization (n = 1,020) from a cohort of adult Americans of European descent participating in the Atherosclerosis Risk in Communities study and employed a MAGENTA approach (i.e., meta-analysis gene set enrichment of variant associations) to obtain gene-centric and gene set association results corrected for gene size, number of single-nucleotide polymorphisms, and local linkage disequilibrium characteristics based on the human genome build 18 (National Center for Biotechnology Information build 36). We used the Gene Ontology, Ingenuity, KEGG, Panther, Reactome, and Biocarta databases for gene set enrichment analyses. Six genes showed evidence of statistically significant association: 4 with severe CP (NIN, p = 1.6 × 10−7; ABHD12B, p = 3.6 × 10−7; WHAMM, p = 1.7 × 10−6; AP3B2, p = 2.2 × 10−6) and 2 with high periodontal pathogen colonization (red complex–KCNK1, p = 3.4 × 10−7; Porphyromonas gingivalis–DAB2IP, p = 1.0 × 10−6). Top-ranked genes for moderate CP were HGD (p = 1.4 × 10−5), ZNF675 (p = 1.5 × 10−5), TNFRSF10C (p = 2.0 × 10−5), and EMR1 (p = 2.0 × 10−5). Loci containing NIN, EMR1, KCNK1, and DAB2IP had showed suggestive evidence of association in the earlier single-nucleotide polymorphism–based analyses, whereas WHAMM and AP2B2 emerged as novel candidates. The top gene sets included severe CP (“endoplasmic reticulum membrane,” “cytochrome P450,” “microsome,” and “oxidation reduction”) and moderate CP (“regulation of gene expression,” “zinc ion binding,” “BMP signaling pathway,” and “ruffle”). Gene-centric analyses offer a promising avenue for efficient interrogation of large-scale GWAS data. These results highlight genes in previously identified loci and new candidate genes and pathways possibly associated with CP, which will need to be validated via replication and mechanistic studies.

Identification of Potential Biomarkers by Serum Proteomics Analysis in Rats with Sepsis

This study was aimed to find new biomarkers for diagnosis and prediction of prognosis of sepsis. Serum samples from nonsurvivor, survivor, and control groups were obtained at 12 h after the induction of sepsis and labeled with isobaric tags (iTRAQ) and then analyzed by two-dimensional liquid chromatography and tandem mass spectrometry. Protein identification and quantification were obtained using mass spectrometry and the ProteinPilot software. Bioinformatics annotation was performed by searching against the PANTHER database. Enzyme-linked immunosorbent assays were used to further confirm the protein identification and differential expression. A logistic regression was then used to screen the index set for diagnosis and prognosis of sepsis. We found that 47 proteins were preferentially elevated in septic rats (both nonsurvivors and survivors) compared with the control rats, and 28 proteins were preferentially elevated in the NS rats as compared with the S group. Several biomarkers, such as multimerin 1, ficolin 1, carboxypeptidase N (CPN2), serine protease 1, and platelet factor 4, were tightly correlated with the diagnosis of sepsis. Logistic regression analyses established multimerin 1, pro-platelet basic protein, fibrinogen-α, and fibrinogen-β for prognosis of sepsis.

Efficiency and Safety of O6-Methylguanine DNA Methyltransferase (MGMTP140K)-MediatedIn VivoSelection in a Humanized Mouse Model

Efficient O⁶-methylguanine DNA methyltransferase (MGMT(P140K))-mediated myeloprotection and in vivo selection have been demonstrated in numerous animal models and most recently in a phase I clinical study in glioblastoma patients. However, this strategy may augment the genotoxic risk of integrating vectors because of chemotherapy-induced DNA damage and the proliferative stress exerted during the in vivo selection. Thus, to improve the safety of the procedure, we evaluated a self-inactivating lentiviral MGMT(P140K) vector for transduction of human cord blood-derived CD34⁺ cells followed by transplantation of the cells into NOD/LtSz-scid/Il2rγ⁻/⁻ mice. These experiments demonstrated significant and stable enrichment of MGMT(P140K) transgenic human cells in the murine peripheral blood and bone marrow. Clonal inventory analysis utilizing linear amplification-mediated polymerase chain reaction and high-throughput sequencing revealed a characteristic lentiviral integration profile. Among the bone marrow insertions retrieved, we observed considerable overlap to previous MGMT(P140K) preclinical models or the clinical study. However, no significant differences between our chemotherapy-treated and nontreated cohorts were observed. This also hold true when specific cancer gene databases and a functional annotation of hit genes by the Panther Database with respect to molecular function, biological process, or cellular component were assessed. Thus, in summary, our data demonstrate efficient and long-term in vivo selection without overt hematological abnormalities using the lentiviral MGMT(P140K) vector. Furthermore, the study introduces humanized mouse models as a novel tool for the pre-clinical assessment of human gene therapy related toxicity.

To the novel paradigm of proteome-based cell therapy of tumors: through comparative proteome mapping of tumor stem cells and tissue-specific stem cells of humans.

We performed proteome mapping (PM), cataloging, and bioinformation analysis of protein lysates of human neural (CD133(+)) progenitor and stem cells (NPSCs) isolated from the olfactory sheath of a nose, multipotent mesenchymal (CD29(+), CD44(+), CD73(+), CD90(+), CD34(-)) stromal cells (MMSCs) isolated from human bone marrow, and tumor (CD133(+)) stem cells (TSCs) isolated from the human U87 glioblastoma (GB) cell line. We identified 1,664 proteins in the examined lysates of stem cells (SCs), 1,052 (63.2%) of which are identical in NPSCs and TSCs and 607 proteins (36.47%) of which are identical in MMSCs and TSCs. Other proteins in U87 GB TSCs are oncospecific or carcinogenesis associated. The biological processes, molecular functions, cell localization, and protein signal pathways of the proteins available in all three proteomes were annotated by PubMed (http://www.ncbi.nlm.nih.gov/pubmed/), PANTHER (http://www.pantherdb.org/), GeneOntology (http://www.geneontology.org/), and KEGG (http://www.genome.jp/kegg/) databases. It was shown that gliomaspheres of U87 GB had only 10 intracellular signal transduction pathways (ISTP) that were not modified by the neoplastic process, but only two of them (integrin and focal adhesion pathways) were accessible for regulatory action on gene candidates in the TSC nucleus. Carcinogenesis-free membrane proteins, IPST, and genes expressing proteins of these pathways in U87 GB TSCs can be viewed as main targets for regulatory effects on TSCs. We offer a novel concept of proteome-based complex therapy of tumors. This manuscript is published as part of the International Association of Neurorestoratology (IANR) special issue of Cell Transplantation.

THU0127 Functional activity of membrane-bound glucocorticoid receptors

BackgroundGlucocorticoids are among the most commonly used anti-inflammatory and immunosuppressive drugs in the treatment of rheumatic diseases. They exert their anti-inflammatory and immunosuppressive effects primarily via the cytosolic glucocorticoid receptor...

Analysis of FOS, BTG2, and NR4A in the function of renal medullary hypertension

The aim of this study was to identify differentially expressed genes (DEGs) in renal medullary hypertension and reveal their pathogenic mechanisms. We downloaded the gene expression profile of GSE28360 from the Gene Expression Omnibus database. The profile included 14 samples (5 normal and 9 hypertension). The DEGs in normal and disease samples were distinguished with a false-discovery rate threshold of <0.05 and a fold-change value of >2 or <-2. We put the selected genes into the online program String 8.3 to obtain the protein-protein interaction network and selected the hub proteins. These hub proteins were then placed in the PANTHER database to determine hub protein-related pathways and explain their functions. Finally, we cleared up the single-nucleotide polymorphisms (SNPs) of the hub genes via combing with the National Center for Biotechnology SNP database. A total of 13 genes were identified as DEGs between normal and disease samples. Five selected hub proteins, B-cell translocation gene 2 (BTG2), FBJ murine osteosarcoma viral oncogene homolog (FOS), nuclear receptor subfamily 4, group A, member 1 (NR4A1), NR4A member 2 (NR4A2), and NR4A member 3 (NR4A3), were mainly related to angiogenesis and B-cell activation. After SNP analysis, 103, 103, 595, 150, and 493 SNPs were found to correspond to BTG2, FOS, NR4A1, NR4A2, and NR4A3, respectively. Our results suggest that pathways of angiogenesis and B-cell activation may involve in the progression of renal medulla hypertension.

Gene expression profiles regulated by Hspa1b in MPTP-induced dopaminergic neurotoxicity using knockout mice

Heat shock proteins 70 (Hsp70), a chaperone that is up-regulated in stress responses and that refolds proteins, might be involved in the pathogenesis of Parkinson disease (PD). This study examined the gene expression profiling regulated by the Hsp70 gene using the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated PD model of mice. The differences in gene expression after a MPTP treatment in C57BL/6 normal mice and Hspa1b (Hsp70-1) knockout (KO) mice was compared using a 24K cDNA microarray. Microarray analysis showed that 246 genes were expressed differentially by the MPTP treatment in normal mice (147 up-regulated and 279 down-regulated). The MPTP-treated KO mice resulted in the differential expression of 26 genes (12 up-regulated and 13 down-regulated) compared to the MPTP-treated normal mice group. The PANTHER database was used to evaluate the microarray data. A total of 38 signaling pathways involved in 426 differentially expressed genes as a result of the MPTP treatment in normal mice were significant. Of these 38 pathways, the pathways related to 25 genes changed by a Hspa1b deficiency were 5-Hydroxytryptamine biosynthesis, adrenaline and noradrenaline biosynthesis, and Parkinson disease. These results demonstrated various genes regulated by Hspa1b in a MPTP-induced PD model. In conclusion, we suggest that this study may be partly helpful to identify action mechanism in the development of a novel drug targeted to Hspa1b gene in the treatment of PD.

Altered protein expression in serum from endometrial hyperplasia and carcinoma patients

BackgroundEndometrial carcinoma is one of the most common gynecological malignancies in women. The diagnosis of the disease at early or premalignant stages is crucial for the patient's prognosis. To date, diagnosis and follow-up of endometrial carcinoma and hyperplasia require invasive procedures. Therefore, there is considerable demand for the identification of biomarkers to allow non-invasive detection of these conditions.MethodsIn this study, we performed a quantitative proteomics analysis on serum samples from simple endometrial hyperplasia, complex endometrial hyperplasia, atypical endometrial hyperplasia, and endometrial carcinoma patients, as well as healthy women. Serum samples were first depleted of high-abundance proteins, labeled with isobaric tags (iTRAQ™), and then analyzed via two-dimensional liquid chromatography and tandem mass spectrometry. Protein identification and quantitation information were acquired by comparing the mass spectrometry data against the International Protein Index Database using ProteinPilot software. Bioinformatics annotation of identified proteins was performed by searching against the PANTHER database.ResultsIn total, 74 proteins were identified and quantified in serum samples from endometrial lesion patients and healthy women. Using a 1.6-fold change as the benchmark, 12 proteins showed significantly altered expression levels in at least one disease group compared with healthy women. Among them, 7 proteins were found, for the first time, to be differentially expressed in atypical endometrial hyperplasia. These proteins are orosomucoid 1, haptoglobin, SERPINC 1, alpha-1-antichymotrypsin, apolipoprotein A-IV, inter-alpha-trypsin inhibitor heavy chain H4, and histidine-rich glycoprotein.ConclusionsThe differentially expressed proteins we discovered in this study may serve as biomarkers in the diagnosis and follow-up of endometrial hyperplasia and endometrial carcinoma.

BioPAX Support in CellDesigner

AbstractBioPAX is a pathway data exchange language that is widely used by pathway database resources. Here we describe a mechanism that allows users to curate pathway data in diagrams using CellDesigner, a pathway graphical editing tool, and save the pathway data in BioPAX format. We used this mechanism to create BioPAX files for all 165 pathways from the PANTHER database. In addition, we are also working on graphically visualizing BioPAX data by importing them into CellDesigner.

Morphinome – A meta‐analysis applied to proteomics studies in morphine dependence

This review is a meta-analysis of data describing proteins regulated by morphine influence studied worldwide across last years administration. Up to date (July 2010), 15 studies concerning this subject have been published. Animal models, examined brain structures, the route of morphine administration and proteomic platforms used for identification of differentially expressed proteins were described. Standardization of obtained results allowed for creation of database of proteins, whose expression was altered by morphine administration (www.addiction-proteomics.org). Their analysis by tools available in Celera Panther Database was possible too. Proteins, which seem to be the most promising candidates for further research, due to their consistent appearance in different studies, were indicated. Created database may facilitate further studies by providing a possibility to compare results obtained during different experiments. At the end, dynamic picture of proteome after morphine administration, which emerges from the obtained results, is discussed and need for standardization of proteomics experiments is stressed. As meta-analysis is a very powerful tool for evaluation and comparison of multiple data. We believe this approach will be useful in the nearest future to extract vital information from a vast number of similar publications. Morphinome database created already by our group is a comfortable tool for validation and verification of new data received after morphine influence on proteomes investigations. It gives a chance for fast comparison of results without hours spent on life science literature mining.

Profiling of hypothalamic and hippocampal gene expression in chronically stressed rats treated with St. John’s wort extract (STW 3-VI) and fluoxetine

Hypericum perforatum L., known as St. John's wort (SJW), is used as a phytotherapeutic agent for the treatment of mild to moderate forms of depression. The aim of the present study was to evaluate the effect of SJW extract (STW 3-VI; 250 and 500 mg/kg; p.o.) and fluoxetine (10 mg/kg, p.o.) on genes involved in the pathogenesis of depression using a chronic restraint stress (CRS) model in rats. Of particular interest was the assessment of similarities and differences between SJW extract and fluoxetine on the gene expression level in two different brain regions. Hypothalamic and hippocampal tissues were analyzed using the Affymetrix gene chip Rat Genome 230 2.0 Array, which comprises more than 30,000 rat transcripts. Limma program and PANTHER database were used to evaluate the microarray data. Genes involved in the pathways of inflammatory processes (Mapk8), oxidative stress (Gpx3, Gstm3, Sod3) or Alzheimer's disease (Sncb, Apbb1ip) were altered by both fluoxetine and SJW extract. For all groups, several signaling pathways were identified which could provide a link between the various hypotheses of depression. In conclusion, microarray analysis proved to be a valuable tool to identify a large number of genes and resulting pathways that may serve as novel drug targets or predict drug responsiveness for SJW or fluoxetine. Based on our comprehensive analysis, it was possible to identify similarities and differences between SJW and fluoxetine which may help to better understand their molecular action and, in addition, help to find novel treatment strategies for stress-related depression.

Microarray Analysis as a Tool to Elucidate the Mechanism of Action of St. John's Wort

Hypericum perforatum L., known as St. John's wort (SJW) is indicated as phytotherapeutic agent for the treatment of mild to moderate forms of depression. Data from literature support the hypothesis that chronic stress is a major risk factor for psychiatric illnesses. The aim of the present study was to evaluate the effect of SJW extract (STW3-VI; 250 and 500mg/kg; p.o.) and fluoxetine (10mg/kg, p.o.) on genes involved in the pathogenesis of depression using a chronic restraint stress (CRS) model in rats. Hypothalamic and hippocampal tissues were analyzed using the Affymetrix gene chip Rat Genome 230 2.0 Array, which comprises more than 30,000 rat transcripts. Limma analysis and PANTHER database were used to evaluate the microarray data. Treatment with SJW extract in both concentrations and fluoxetine reversed expression levels of several genes changed by CRS. Genes involved in pathways of inflammatory processes (Mapk8) oxidative stress (Gpx3, Gstm3, Sod3) or Alzheimer's disease (Sncb, Apbb1ip) were altered by both, fluoxetine and SJW. In all groups several pathways were identified which provide a link between the various hypotheses of depression. Gene expression results for both brain regions were verified using quantitative rPCR. In conclusion, Microarray analysis prooved to be a valuable tool to identify a large number of genes and resulting pathways that may serve as novel drug targets or predict drug responsiveness. Based on our results it was possible to identify new candidate pathways that help to understand molecular principles of depression and in addition help to find novel treatment strategies for neuropsychiatric disorders.

Transcriptome Analysis Describing New Immunity and Defense Genes in Peripheral Blood Mononuclear Cells of Rheumatoid Arthritis Patients

BackgroundLarge-scale gene expression profiling of peripheral blood mononuclear cells from Rheumatoid Arthritis (RA) patients could provide a molecular description that reflects the contribution of diverse cellular responses associated with this disease. The aim of our study was to identify peripheral blood gene expression profiles for RA patients, using Illumina technology, to gain insights into RA molecular mechanisms.Methodology/Principal FindingsThe Illumina Human-6v2 Expression BeadChips were used for a complete genome-wide transcript profiling of peripheral blood mononuclear cells (PBMCs) from 18 RA patients and 15 controls. Differential analysis per gene was performed with one-way analysis of variance (ANOVA) and P values were adjusted to control the False Discovery Rate (FDR<5%). Genes differentially expressed at significant level between patients and controls were analyzed using Gene Ontology (GO) in the PANTHER database to identify biological processes. A differentially expression of 339 Reference Sequence genes (238 down-regulated and 101 up-regulated) between the two groups was observed. We identified a remarkably elevated expression of a spectrum of genes involved in Immunity and Defense in PBMCs of RA patients compared to controls. This result is confirmed by GO analysis, suggesting that these genes could be activated systemically in RA. No significant down-regulated ontology groups were found. Microarray data were validated by real time PCR in a set of nine genes showing a high degree of correlation.Conclusions/SignificanceOur study highlighted several new genes that could contribute in the identification of innovative clinical biomarkers for diagnostic procedures and therapeutic interventions.