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At least 1,495 records · Page 83Linked to original sources
PubMed2006

Differential gene expression profiles between tumor biopsies and short-term primary cultures of ovarian serous carcinomas: identification of novel molecular biomarkers for early diagnosis and therapy.

OBJECTIVE: To identify novel molecular biomarkers useful for the early diagnosis and therapy of ovarian cancer by gene expression profiling. To compare the genetic fingerprints of flash-frozen ovarian serous carcinomas to those of matched highly purified primary tumor cell cultures. METHODS: Gene expression profiles of 19 flash-frozen ovarian serous papillary carcinoma (OSPC) were analyzed and compared to 15 controls (highly purified human ovarian surface epithelium short-term cultures, HOSE) using oligonucleotide microarrays complementary to >14,500 human genes. In addition, gene expression profiling of 5 highly purified primary OSPC cultured in vitro for less than 2 weeks was compared to flash-frozen ovarian carcinoma biopsies obtained from matched samples. Quantitative RT-PCR and IHC staining techniques were used to validate microarray data at RNA and protein levels for some of the differentially expressed genes. RESULTS: Unsupervised analysis of gene expression data readily distinguished normal tissue from flash-frozen OSPC and identified 901 and 557 genes that exhibited >3-fold up-regulation or down-regulation, respectively, in OSPC when compared to HOSE. Mammaglobin 2, an ovarian secreted protein, was identified as the top differentially expressed gene in OSPC (19 out 19 OSPC versus 0 out of 15 HOSE) with over 827-fold up-regulation relative to HOSE. The claudin and kallikrein family of proteins including the clostridium perfringens enterotoxin receptors claudin 3 and 4, kallikreins 6, 7, 8, 10, 11 and the immunomodulatory molecule B7-H4 were found among the most highly overexpressed genes in OSPC when compared to HOSE. Genetic fingerprints of flash-frozen OSPC were found to have high correlation with those of purified primary OSPC short-term in vitro cultures with only 31 out of 8,637 genes (0.35%) differentially expressed between the two groups. CONCLUSIONS: Short-term in vitro culture of primary ovarian carcinomas may greatly increase the purity of ovarian tumor RNA available for gene expression profiling without causing major alteration in OSPC fingerprints. Mammaglobin 2, kallikreins 6, 7, 8, 10, 11, claudin 3 and 4 and B7-H4 gene expression products represent candidate biomarkers endowed with great potential for early screening and therapy of OSPC patients.

Adult↗
PubMed2002

Gene expression profiling of mouse bladder inflammatory responses to LPS, substance P, and antigen-stimulation.

Inflammatory bladder disorders such as interstitial cystitis (IC) deserve attention since a major problem of the disease is diagnosis. IC affects millions of women and is characterized by severe pain, increased frequency of micturition, and chronic inflammation. Characterizing the molecular fingerprint (gene profile) of IC will help elucidate the mechanisms involved and suggest further approaches for therapeutic intervention. Therefore, in the present study we used established animal models of cystitis to determine the time course of bladder inflammatory responses to antigen, Escherichia coli lipopolysaccharide (LPS), and substance P (SP) by morphological analysis and cDNA microarrays. The specific aim of the present study was to compare bladder inflammatory responses to antigen, LPS, and SP by morphological analysis and cDNA microarray profiling to determine whether bladder responses to inflammation elicit a specific universal gene expression response regardless of the stimulating agent. During acute bladder inflammation, there was a predominant infiltrate of polymorphonuclear neutrophils into the bladder. Time-course studies identified early, intermediate, and late genes that were commonly up-regulated by all three stimuli. These genes included: phosphodiesterase 1C, cAMP-dependent protein kinase, iNOS, beta-NGF, proenkephalin B and orphanin, corticotrophin-releasing factor (CRF) R, estrogen R, PAI2, and protease inhibitor 17, NFkB p105, c-fos, fos-B, basic transcription factors, and cytoskeleton and motility proteins. Another cluster indicated genes that were commonly down-regulated by all three stimuli and included HSF2, NF-kappa B p65, ICE, IGF-II and FGF-7, MMP2, MMP14, and presenilin 2. Furthermore, we determined gene profiles that identify the transition between acute and chronic inflammation. During chronic inflammation, the urinary bladder presented a predominance of monocyte/macrophage infiltrate and a concomitant increase in the expression of the following genes: 5-HT 1c, 5-HTR7, beta 2 adrenergic receptor, c-Fgr, collagen 10 alpha 1, mast cell factor, melanocyte-specific gene 2, neural cell adhesion molecule 2, potassium inwardly-rectifying channel, prostaglandin F receptor, and RXR-beta cis-11-retinoic acid receptor. We conclude that microarray analysis of genes expressed in the bladder during experimental inflammation may be predictive of outcome. Further characterization of the inflammation-induced gene expression profiles obtained here may identify novel biomarkers and shed light into the etiology of cystitis.

Animals↗
PubMed2004

Quantitative gene expression profiling in formalin-fixed, paraffin-embedded tissues using universal bead arrays.

We recently developed a sensitive and flexible gene expression profiling system that is not dependent on an intact poly-A tail and showed that it could be used to analyze degraded RNA samples. We hypothesized that the DASL (cDNA-mediated annealing, selection, extension and ligation) assay might be suitable for the analysis of formalin-fixed, paraffin-embedded tissues, an important source of archival tissue material. We now show that, using the DASL assay system, highly reproducible tissue- and cancer-specific gene expression profiles can be obtained with as little as 50 ng of total RNA isolated from formalin-fixed tissues that had been stored from 1 to over 10 years. Further, tissue- and cancer-specific markers derived from previous genome-wide expression profiling studies of fresh-frozen samples were validated in the formalin-fixed samples. The DASL assay system should prove useful for high-throughput expression profiling of archived clinical samples.

Algorithms↗
PubMed2003

Gene expression profiling of alcoholic liver disease in the baboon (Papio hamadryas) and human liver.

The molecular pathogenesis of alcoholic liver disease (ALD) is not well understood. Gene expression profiling has the potential to identify new pathways and altered molecules in ALD. Gene expression profiles of ALD in a baboon model and humans were compared using DNA arrays. Reverse transcriptase-polymerase chain reaction and immunohistochemistry were used for downstream analysis of array results. cDNA array analysis revealed differential expression of several novel genes and pathways in addition to genes known to be involved in ALD pathogenesis. Overall gene expression profiles were similar in both species, with a majority of genes involved with fibrogenesis and xenobiotic metabolism, as well as inflammation, oxidant stress, and cell signaling. Genes associated with stellate cell activation (collagens, matrix metalloproteinases, tissue inhibitors of matrix metalloproteinase) were up-regulated in humans. Decreased expression of several metallothioneins was unexpected. Fourteen molecules related to the annexin family were up-regulated, including annexin A1 and A2. Immunofluorescence revealed a marked overexpression of annexin A2 in proliferating bile duct cells, hepatocyte cell surface, and selective co-localization with CD14-positive cells in human ALD. The gene expression profile of ALD is dominated by alcohol metabolism and inflammation and differs from other liver diseases. Annexins may play a role in the progression of fibrosis in ALD.

Adult↗
PubMed2002

Molecular characterization of human meningiomas by gene expression profiling using high-density oligonucleotide microarrays.

Meningiomas are common central nervous system neoplasms that exhibit remarkably diverse histopathology and biological behavior. Compared to astrocytomas, the most common central nervous system tumor, little is known about the molecular pathways critical for meningioma tumor formation and malignant progression. As an initial step toward characterizing the genetic basis of meningioma pathogenesis, we assessed cancer-related gene expression profiles of nonneoplastic leptomeningeal specimens and human meningiomas of varying World Health Organization (WHO) grade using high-density oligonucleotide microarrays. Although expression profile differences between nonneoplastic and meningioma specimens were readily discernible, the expression profile of a subset of genes could also distinguish WHO grade I from WHO grades II and III tumors. Altered expression levels of several genes identified in this study have been previously noted in meningiomas (eg, growth hormone receptor, IGFBP-7, endothelin receptor A, IGF2). However, we also identified a number of novel genes whose expression was associated with WHO grade and was confirmed by reverse transcriptase-polymerase chain reaction in a larger, independent set of meningeal tumors (n = 47). This report represents the first gene expression profiling studies of meningiomas and identifies some initial candidate genes that may provide further insights into the genetic basis for meningioma pathogenesis.

Carrier Proteins↗
PubMed2002

mRNA expression profiling of laser microbeam microdissected cells from slender embryonic structures.

Microarray hybridization has rapidly evolved as an important tool for genomic studies and studies of gene regulation at the transcriptome level. Expression profiles from homogenous samples such as yeast and mammalian cell cultures are currently extending our understanding of biology, whereas analyses of multicellular organisms are more difficult because of tissue complexity. The combination of laser microdissection, RNA amplification, and microarray hybridization has the potential to provide expression profiles from selected populations of cells in vivo. In this article, we present and evaluate an experimental procedure for global gene expression analysis of slender embryonic structures using laser microbeam microdissection and laser pressure catapulting. As a proof of principle, expression profiles from 1000 cells in the mouse embryonic (E9.5) dorsal aorta were generated and compared with profiles for captured mesenchymal cells located one cell diameter further away from the aortic lumen. A number of genes were overexpressed in the aorta, including 11 previously known markers for blood vessels. Among the blood vessel markers were endoglin, tie-2, PDGFB, and integrin-beta1, that are important regulators of blood vessel formation. This demonstrates that microarray analysis of laser microbeam micro-dissected cells is sufficiently sensitive for identifying genes with regulative functions.

Animals↗
PubMed2002

DNA microarray analysis of the expression profiles of luteinized granulosa cells as a function of ovarian reserve.

OBJECTIVE: To examine the expression profiles of luteinized granulosa cells isolated from women with normal or diminished ovarian reserve undergoing in vitro fertilization. DESIGN: Expression profiling by complementary DNA microarray analysis. SETTING: Women undergoing in vitro fertilization-embryo transfer in a university-based fertility clinic. PATIENT(S): Eighteen women with normal or decreased ovarian reserve. INTERVENTION(S): All patients were given gonadotropin stimulation in preparation for IVF with granulosa cells isolated at the time of follicular aspiration. MAIN OUTCOME MEASURE(S): Expression profiles of luteinized granulosa cells isolated from each woman were determined by using DNA microarray analysis. RESULT(S): Changes in patterns of gene expression in granulosa cells were observed between women with normal and diminished ovarian reserve. These genes included several anonymous expressed sequence tags and also expressed sequence tags that correspond to known genes. CONCLUSION(S): Expression profiling of granulosa cells by DNA microarray may yield signature patterns that reflect the status of ovarian reserve and the competence of granulosa cells.

Adult↗
PubMed2000

Gene expression profiling in human peripheral blood mononuclear cells using high-density filter-based cDNA microarrays.

Microarray technology has provided the ability to analyse the expression profiles for thousands of genes in parallel. The need for highly specialised equipment to use certain types of microarrays has restricted the application of this technology to a small number of dedicated laboratories. High-density filter-based cDNA microarrays provide a low-cost option for performing high-throughput gene expression analysis. We have used a model system in which filter-based cDNA microarrays representing over 4000 known human genes were used to monitor the kinetics of gene expression in human peripheral blood mononuclear cells (PBMCs) stimulated with phytohaemagluttinin (PHA). Using software-based cluster analysis, we identified 104 genes that altered in expression levels in response to PHA stimulation of PBMCs and showed that there was a considerable overlap between genes with similar temporal expression profiles and similar functional roles. Comparison of microarray quantitation with quantitative PCR showed almost identical expression profiles for a number of genes. Coupled with the fact that our findings are in agreement with a large number of independent observations, we conclude that the use of filter-based cDNA microarrays is a valid and accurate method for high-throughput gene expression profiling.

Cells, Cultured↗
PubMed2003

Transcriptional profiling in hepatoblastomas using high-density oligonucleotide DNA array.

Hepatoblastoma is a common hepatic tumor in children. Although evidence regarding cytogenetic and molecular genetic alterations in hepatoblastomas has been reported, the molecular events affecting the biologic characteristics of this tumor, including alterations of the gene expression profile, are largely unknown. To identify genes differentially expressed between nondiseased liver (NDL) and hepatoblastoma tumor (HBT), we analyzed the gene expression profile in 14 NDL and 16 HBT samples using a high-density oligonucleotide DNA array. Using Mann-Whitney U test followed by the k-nearest neighbor algorithm, we identified 26 genes (predictor genes) that were able to assign unknown samples derived from NDL and HBT to either the NDL group or HBT group with 100% accuracy. Using a cross-validation approach, we confirmed that the k-nearest neighbor algorithm assigned the particular samples derived from NDL and HBT to either the NDL or HBT group with 93.3% (28/30 samples) accuracy. In the 26 predictor genes, we found alteration of the expression of genes regulating cell division (NAP1L1, STMN1, CCNG2, and CDC7L1) and tumor cell growth (IGF2 and IGFBP4) in HBT. Four predictor genes (ETV3, TPR, CD34, and NR1I3) were also found to be mapped to the chromosomal region 1q21 approximately q32, which has been reported to be frequently involved in the development of hepatoblastoma. The findings obtained in this study suggest that alteration of the expression of some genes regulating cell division and tumor cell growth may be characteristics of the gene expression profile in HBT, and that alteration of the expression of the four predictor genes mapped to chromosomal region 1q21 approximately q32 may also contribute to the differences in gene expression profile between NDL and HBT.

Child↗
PubMed2001

Expression profile of active genes in human periodontal ligament and isolation of PLAP-1, a novel SLRP family gene.

Periodontal ligament (PDL) is one of the most important tissues in maintaining the homeostasis of tooth and tooth-supporting tissue, periodontium. In this study, we investigated the expression profile of active genes in the human PDL obtained by collecting sequences with a 3'-directed cDNA library, which faithfully represents the composition of the mRNA population. We succeeded in obtaining a total of 1752 cDNA sequences by sequencing randomly selected clones and identified a total of 1318 different species as gene signatures (GS) by their sequence identity, 344 of which were known genes in the GenBank, and 974 of which were new genes. The resulting expression profile showed that collagen type I and type III were the most abundant genes and that osteogenesis-related proteins, such as SPARC/osteonectin and osteoblast specific factor 2, were highly expressed. By comparing the expression profile of PDL with 44 profiles similarly obtained with unrelated human cell/tissue, nine novel genes, which are probably expressed specifically in PDL, were discovered. Among them, we cloned a full-length cDNA of GS5096, which is frequently expressed in freshly-isolated periodontal tissue. We found that it encodes a novel protein, which is a new member of the class I small leucine-rich repeat proteoglycan family, and designated it PLAP-1 (periodontal ligament associated protein-1). PLAP-1 mRNA expression was confirmed in in vitro-maintained PDL cells and was enhanced during the course of the cytodifferentiation of the PDL cells into mineralized tissue-forming cells such as osteoblasts and cementoblasts. These findings suggest the involvement of PLAP-1 in the mineralized matrix formation in PDL tissues.

Amino Acid Sequence↗
PubMed2001

Gene expression profiling in the post-mortem human brain--no cause for dismay.

Global expression profiling techniques such as microarray technology promise to revolutionize biology. Soon it will be possible to investigate alterations at the transcript level of the entire human genome. There is great hope that these techniques will at last shed light on the pathological processes involved in complex neuropsychiatric disorders such as schizophrenia. These scientific advances in turn have re-kindled a great interest and demand for post-mortem brain tissue. Good quality post-mortem tissue undoubtedly is the fundamental prerequisite to investigate complex brain disorders with molecular profiling techniques. In this review we show that post-mortem brain tissue can yield good quality mRNA and intact protein antigens which allow the successful application of traditional molecular biology methods as well as novel profiling techniques. We also consider the use of laser-capture microdissection on post-mortem tissue. This recently developed technique allows the experimenter to explore the molecular basis of cellular function at the single cell level. The combination of laser-capture microdissection with high throughput profiling techniques offers opportunities to obtain precise genetic fingerprints of individual neurons allowing comparisons of normal and pathological states.

Autopsy↗
PubMed2003

Accurate and reproducible gene expression profiles from laser capture microdissection, transcript amplification, and high density oligonucleotide microarray analysis.

Gene expression profiling using high density oligonucleotide arrays is a powerful method to generate an unbiased survey of a cell's transcriptional landscape. Increasingly complex biological questions require that this approach be applicable to the small numbers of cells that are obtained from sources such as laser capture microdissection (LCM) of solid tissues. In this report, we demonstrate that two rounds of transcript amplification can generate accurate and reproducible gene expression profiles using high density oligonucleotide microarrays, starting with as little as 10 ng of total RNA. Biased amplification of the 3' end of transcripts does not have a major impact on the overall transcript profile due to the 3' bias of probe sets incorporated in the array design. Furthermore, greater than 95% of all genes detected demonstrate less than a twofold difference in expression when independent tissue dissections of identical cell populations are compared. The accuracy and technical reproducibility of the method suggests that expression profiling using transcript amplification and high density oligonucleotide microarrays can be used on a routine basis.

Breast Neoplasms↗
PubMed2002

Core biopsies can be used to distinguish differences in expression profiling by cDNA microarrays.

The primary focus of this work was to determine the feasibility of obtaining representative expression array profiles from clinical core biopsies. For this purpose we performed six 16-gauge needle core biopsies and an excision biopsy on each of two different human xenografts, one from an Ewing's sarcoma cell line and the second from neuroblastoma cell line grown in Beige-Scid mice. Three of the six core biopsies were processed separately and the remaining three were pooled and processed together. As the initial RNA material isolated from the core biopsies was not sufficient for microarray analysis, an amplification procedure using a modified Eberwine protocol was performed, and the amplified products applied onto a 6000-feature human cDNA microarray. Comparisons of the array results from core biopsies (amplified RNA) and surgical specimens (non-amplified RNA) showed maintenance of the expression profile as assessed by hierarchical clustering. Gene expression profiles obtained from microarray analysis clearly differentiated the Ewing's sarcoma from the neuroblastoma with both core and excisional biopsies as starting material. Pooling the core biopsies did not improve the concordance with excisional biopsies. In conclusion, our results suggest that core biopsies can be used as a suitable and reliable material for the determination of tumor genetic profiles.

Animals↗
PubMed2002

Expression profiling to understand actions of NMDA/glutamate receptor antagonists in rat brain.

Agents acting as noncompetitive N-methyl-D-aspartate (NMDA)/glutamate receptor antagonists induce the expression of several genes in limbic cortical regions, such as the cingulate, retrosplenial, and entorhinal cortices. These include important regulatory genes such as the neurotrophin brain-derived neurotrophic factor (BDNF), its receptor trkB, and c-fos. We applied expression profiling methods to find genes coregulated with BDNF following treatment with the prototypical NMDA/glutamate receptor antagonist MK-801. Expression profiling provides a useful technique for describing the molecular and transcriptional level events that follow various processes. We illustrate the utility of microarrays to find novel ESTs regulated by MK-801. We also used expression profiling with microarrays to characterize the levels of transcription factor cAMP response element modulator (CREM) and inducible cAMP early repressor (ICER) isoforms that are induced by MK-801. These factors may act as the eventual repressors for BDNF expression via competition and heterodimerization with phosphorylated CREB, a transcription factor important for BDNF expression. Finally, we find and confirm the regulation of Erp29, RTNI, and an ABC transporter by antagonism of NMDA/glutamate receptors as potential stress related molecules in brain. The emerging picture generated by using these expression profiling approaches, identifies several of what likely will be many molecules that take part in the complex events that occur during BDNF signaling mediated by blockade of NMDA/ glutamate receptors.

Animals↗
PubMed2003

Gene expression profiling in ecotoxicology.

Gene expression profiling is a powerful new end point for ecotoxicology and a means for bringing the genomics revolution to this field. We review the usefulness of gene expression profiling as an end point in ecotoxicology and describe methods for applying this approach to non-model organisms. Since genomes contain thousands of genes representing hundreds of pathways, it is possible to identify toxicant-specific responses from this wide array of possibilities. Stressor-specific signatures in gene expression profiles can be used to diagnose which stressors are impacting populations in the field. Screening for stress-induced genes requires special techniques in organisms without sequenced genomes. These techniques include differential display polymerase chain reaction (DD PCR), suppressive subtractive hybridization PCR (SSH PCR), and representational difference analysis. Gene expression profiling in model organisms like yeast has identified hundreds of genes that are up-regulated in response to various stressors, including several that are well characterized (e.g., hsp78, metallothionein, superoxide dismutase). Using consensus PCR primers from several animal sequences, it is possible to amplify some of these well characterized stress-induced genes from organisms of interest in ecotoxicology. We describe how several stress-induced genes can be grouped into cDNA arrays for rapidly screening samples.

Animals↗
PubMed2004

Expression profile analysis of neurodegenerative disease: advances in specificity and resolution.

Microarray technology has become a common tool for developing expression profiles. Initially used in the analysis of cells lines and homogeneous tissues, this platform has been applied to more diverse tissues, such as the brain. Several neural disorders have already been profiled by microarrays using relatively large amounts of tissue. This data has unveiled many genes with differential expression between normal and diseased tissue that could potentially be used as gene markers for these afflictions. Because of the heterogeneity of the CNS, it is likely that small differences between gene expression in these studies would be enhanced by the sampling of a subset of cells based on these newly characterized gene markers. Subtraction of normal, unaffected cells from the sample may also result in a more accurate profile of a diseased cell. Expression profile studies from several neuropathological states are presented, with emphasis placed on those studies using small samples of cellular material and those using specialized methods of cell isolation and RNA amplification.

AIDS Dementia Complex↗
PubMed2004

Development of the Short Form Endometriosis Health Profile Questionnaire: the EHP-5.

OBJECTIVE: To develop and validate a short form version of the Endometriosis Health Profile-30 questionnaire which consists of a 30-item core questionnaire and a 23 item modular questionnaire. METHODS: Three studies were carried out to develop the Endometriosis Health Profile-5. Study 1: a short form version of the Endometriosis Health Profile-30 core questionnaire was developed. Study 2: the results were verified in a new data set. Study 3: a short form version of the 23 item modular questionnaire was produced. The modular questionnaire contains six dimensions which may not be applicable to every woman with endometriosis and is used to supplement the five scales on the core questionnaire when required. RESULTS: The final instrument contained 11 items: five items from the core questionnaire and six items from the modular questionnaire. The 95% confidence intervals for Study 1 indicated that the scale scores overlapped with each item score and were confirmed in the second analysis (study 2). Each item was most highly correlated to its parent scale and less with the remaining four core questionnaire scales which was verified in study 2. In both studies all correlations were significant at the 0.01 level (two tailed test). CONCLUSION: The Endometriosis Health Profile-5 is a reliable and valid short form questionnaire which can also be supplemented with a short form version of the modular questionnaire when required. It will be especially useful in clinical settings where a short and economical endometriosis health status measure is required.

Adolescent↗
PubMed2004

Expression profiling using a hexamer-based universal microarray.

We describe a transcriptional analysis platform consisting of a universal micro-array system (UMAS) combined with an enzymatic manipulation step that is capable of generating expression profiles from any organism without requiring a priori species-specific knowledge of transcript sequences. The transcriptome is converted to cDNA and processed with restriction endonucleases to generate low-complexity pools (approximately 80-120) of equal length DNA fragments. The resulting material is amplified and detected with the UMAS system, comprising all possible 4,096 (4(6)) DNA hexamers. Ligation to the arrays yields thousands of 14-mer sequence tags. The compendium of signals from all pools in the array-of-universal arrays comprises a full-transcriptome expression profile. The technology was validated by analysis of the galactose response of Saccharomyces cerevisiae, and the resulting profiles showed excellent agreement with the literature and real-time PCR assays. The technology was also used to demonstrate expression profiling from a hybrid organism in a proof-of-concept experiment where a T-cell receptor gene was expressed in yeast.

3' Untranslated Regions↗