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Results for “Tissue Array Analysis”

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At least 253 records · Page 14Linked to original sources
PubMed2002

Altered expression of novel genes in the cerebral cortex following experimental brain injury.

Damage to the cerebral cortex results in neurological impairments such as motor, attention, memory and executive dysfunctions. To examine the molecular mechanisms contributing to these deficits, mRNA expression was profiled using high-density cDNA microarray hybridization after experimental cortical impact injury in mice. The mRNA levels at 2 h, 6 h, 24 h, 3 days and 14 days after injury were compared with those of control animals. This revealed 86 annotated genes and 24 expression sequence tags (ESTs) as being differentially expressed with a 1.5-fold or greater change. Quantitative real-time PCR analysis was used to independently verify these results for selected genes. Seven functional classes of genes were found to be altered following injury, including transcription factors, signal transduction genes and inflammatory proteins. While a few of these genes have been previously reported to be differentially regulated following injury, the most of the genes have not been previously implicated in traumatic brain injury (TBI) pathophysiology. For example, consistent with previous reports, the transcription factor c-jun and the neurotrophic factor bdnf mRNA levels were altered as a result of TBI. Among the novel genes, the mRNA levels for the high mobility group protein 1 (hmg-1), the regulator of G-protein signaling 2 (rgs-2), the transforming growth factor beta inducible early growth response (tieg), the inhibitor of DNA binding 3 (id3), and the heterogeneous nuclear ribonucleoprotein H (hnrnp h) were changed following injury. The functional significance of these genes in neurite outgrowth, neuronal regeneration, and plasticity following injury are discussed.

Animals↗
PubMed2002

A regression-based method to identify differentially expressed genes in microarray time course studies and its application in an inducible Huntington's disease transgenic model.

Time-course studies with microarray technologies provide enormous potential for exploring underlying mechanisms of biological phenomena in many areas of biomedical research, but the large amount of gene expression data generated by such studies also presents great challenges to data analysis. Here we introduce a regression-based statistical modeling approach that identifies differentially expressed genes in microarray time-course studies. To illustrate this method, we applied it to data generated from an inducible Huntington's disease transgenic model. The regression method accounts for the induction process, incorporates relevant experimental information, and includes parameters that specifically address the research interest: the temporal differences in gene expression profiles between the mutant and control mice over the time course, in addition to heterogeneities that commonly exist in microarray data. Least-squares and estimating equation techniques were used to estimate parameters and variances, and inferences were made based on efficient and robust Z-statistics under a set of well-defined assumptions. A permutation test was also used to estimate the number of false-positives, providing an alternative measurement of statistical significance useful for investigators to make decisions on follow-up studies.

Animals↗
PubMed2001

Identification of differentially expressed genes in the denervated rat hippocampus by cDNA arrays.

To elucidate the molecular mechanism underlying the physiological responses to injury in the central nervous system, gene expression profiles in rodent hippocampus following perforant path transection were investigated using cDNA array hybridization. Of the 8000 arrayed clones, 47 exhibited differential expression by >3-fold difference in the denervated hippocampus from control, with 15 up-regulated and 22 down-regulated. They can be functionally assigned into several classes, among which the most prominent are those coding proteins involved in macromolecules synthesis and processing. Northern blot analysis verified the validation of the aforementioned array data. These results throw some new light on the physiological responses of the hippocampus to entorhinal deafferentation at molecular level.

Acetylcholinesterase↗
PubMed2001

EEG1, a putative transporter expressed during epithelial organogenesis: comparison with embryonic transporter expression during nephrogenesis.

A screen for genes differentially regulated in a model of kidney development identified the novel gene embryonic epithelia gene 1 (EEG1). EEG1 exists as two transcripts of 2.4 and 3.5 kb that are most highly expressed at embryonic day 7 and later in the fetal liver, lung, placenta, and kidney. The EEG1 gene is composed of 14 exons spanning a 20-kb region at human chromosome 11p12 and the syntenic region of mouse chromosome 2. Six EEG1 exons have previously been assigned to a longer isoform of eosinophil major basic protein termed proteoglycan 2. Another gene distantly related to EEG1, POV1/PB39, is located 88 kb upstream from the EEG1 gene on chromosome 11. Temporal expression of 65 members of the solute carrier (SLC)-class of transport proteins was followed during kidney development using DNA arrays. POV-1 and EEG1, like glucose transporters, displayed very early maximal gene expression. In contrast, other SLC genes, such as organic anion and cation transporters, amino acid permeases, and nucleoside transporters, had maximal expression later in development. Thus, although the bulk of transporters are expressed late in kidney development, a fraction are expressed near the onset of nephrogenesis. The data raise the possibility that EEG1 and POV1 may define a new family of transport proteins involved in the transport of nutrients or metabolites in rapidly growing and/or developing tissues.

Amino Acid Transport System y+L↗
PubMedDate not supplied

Determination of nitroimidazole residues in poultry tissues, serum and eggs by high-performance liquid chromatography.

A simple high-performance liquid chromatographic (HPLC) method was developed for determination of dimetridazole and metronidazole residues in poultry muscle, liver, serum and eggs. The drug residues were extracted from tissues and egg samples with acetonitrile followed by solid-phase extraction clean-up and HPLC analysis with photodiode array detector. Serum samples were treated with trichloroacetic acid, centrifuged and analysed without further clean-up. The detection limits were 2 ng/g and 5 ng/mL of analysed tissue (egg) and serum samples, respectively. Average recoveries for spike levels of 10 and 20 ng/g ranged from 82 to 94%, and coefficients of variation were below 10%.

Animals↗
PubMed2003

The impact of parietal cells on Helicobacter pylori tropism and host pathology: an analysis using gnotobiotic normal and transgenic mice.

Helicobacter pylori infection of the human stomach is common and typically benign, although a subset of hosts develops severe pathology. Infection occurs in an organ with distinct microenvironments characterized by pronounced differences in the composition of acid-producing parietal cells. In this study, we examine determinants of bacterial tropism to various gastric niches by using germ-free normal and transgenic mice with an engineered parietal cell ablation. Mice were colonized for 8 weeks with a clinical isolate (Hp1) that expresses adhesins recognized by epithelial NeuAcalpha2,3Galbeta1,4 glycan receptors. In normal mice, Hp1 has tropism for a parietal cell-deficient niche where sialylated glycans are expressed by a narrow band of pit cells positioned at the boundary between the squamous epithelium (forestomach) and the proximal glandular epithelium. Lymphoid aggregates that develop in this niche, but not elsewhere in the stomach, were analyzed by GeneChip and quantitative RT-PCR studies of laser capture microdissected mucosa and yielded a series of biomarkers indicative of immune cell activation and maturation. Genetic ablation of parietal cells produced a new source of NeuAcalpha2,3Galbeta1,4 glycans in amplified gastric epithelial lineage progenitors, with accompanying expansion of Hp1 within the glandular epithelium. Lymphoid aggregates that develop in this formerly acid-protected epithelium have molecular features similar to those observed at the forestomach/glandular junction. These findings demonstrate the important roles played by parietal cells and glycan receptors in determining the positioning of H. pylori within the gastric ecosystem, and emphasize the need to consider the evolution of pathology within a given host in a niche-specific context.

Animals↗
PubMed2001

Preprocessing implementation for microarray (PRIM): an efficient method for processing cDNA microarray data.

cDNA microarray technology is useful for systematically analyzing the expression profiles of thousands of genes at once. Although many useful results inferred by using this technology and a hierarchical clustering method for statistical analysis have been confirmed using other methods, there are still questions about the reproducibility of the data. We have therefore developed a data processing method that very efficiently extracts reproducible data from the result of duplicate experiments. It is designed to automatically filter the raw results obtained from cDNA microarray image-analysis software. We optimize the threshold value for filtering the data by using the product of N and R, where N is the ratio of the number of spots that passed the filtering vs. the total number of spots, and R is the correlation coefficient for results obtained in the duplicate experiments. Using this method to process mouse tissue expression profile data that contain 1,881,600 points of analysis, we obtained clustered results more reasonable than those obtained using previously reported filtering methods.

Animals↗
PubMed1991

Ultrasonic tissue characterization of chronic liver disease using cepstral analysis.

To evaluate the morphological changes in chronic liver disease by means of ultrasonography, cepstral analysis was applied to the ultrasonic waveforms obtained with a 10-MHz transducer during peritoneoscopy. Assuming that liver tissue was a collection of semiregularly arrayed small scatters of ultrasound, the spaces between scatterers of the liver could be measured by means of cepstral analysis. From the distribution of the spaces between scatterers, two statistical parameters, mode and kurtosis, were determined for each patient. The mode of spaces between scatterers for liver cirrhosis was significantly larger than that for chronic hepatitis and nonspecific change. The mode of spaces between scatterers was correlated with the size of hepatic lobules or nodules. The kurtosis of spaces between scatterers for chronic hepatitis and liver cirrhosis was significantly smaller than that for nonspecific change. As a preliminary study for noninvasive application of this technique, cepstral analysis was also applied to the ultrasonogram obtained through the abdominal wall with a 3.5-MHz transducer. The results obtained with a 3.5-MHz transducer were correlated well with those obtained with a 10-MHz transducer. These results suggest the possibility of noninvasive evaluation of the structural changes in chronic liver disease.

Chronic Disease↗
PubMed2000

Fatty acid synthesis: from CO2 to functional genomics.

For over 25 years there has been uncertainty over the pathway from CO(2) to acetyl-CoA in chloroplasts. On the one hand, free acetate is the most effective substrate for fatty acid synthesis by isolated chloroplasts, and free acetate concentrations reported in leaf tissue (0.1-1 mM) appear adequate to saturate fatty acid synthase. On the other hand, a clear mechanism to generate sufficient free acetate for fatty acid synthesis is not established and direct production of acetyl-CoA from pyruvate by a plastid pyruvate dehydrogenase seems a more simple and direct path. We have re-examined this question and attempted to distinguish between the alternatives. The kinetics of (13)CO(2) and (14)CO(2) movement into fatty acids and the absolute rate of fatty acid synthesis in leaves was determined in light and dark. Because administered (14)C appears in fatty acids within < 2-3 min our results are inconsistent with a large pool of free acetate as an intermediate in leaf fatty acid synthesis. In addition, these studies provide an estimate of the turnover rate of fatty acid in leaves. Studies similar to the above are more complex in seeds, and some questions about the regulation of plant lipid metabolism seem difficult to solve using conventional biochemical or molecular approaches. For example, we have little understanding of why or how some seeds produce >50% oil whereas other seeds store largely carbohydrate or protein. Major control over complex plant biochemical pathways may only become possible by understanding regulatory networks which provide 'global' control over these pathways. To begin to discover such networks and provide a broad analysis of gene expression in developing oilseeds, we have produced microarrays that display approx. 5000 seed-expressed Arabidopsis genes. Sensitivity of the arrays was 1-2 copies of mRNA/cell. The arrays have been hybridized with probes derived from seeds, leaves and roots, and analysis of expression ratios between the different tissues has allowed the tissue-specific expression patterns of many hundreds of genes to be described for the first time. Approx. 10% of the genes were expressed at ratios > or = 10-fold higher in seeds than in leaves or roots. Included in this list are a large number of proteins of unknown function, and potential regulatory factors such as protein kinases, phosphatases and transcription factors. The arrays were also found to be useful for analysis of Brassica seeds.

Acetyl Coenzyme A↗
PubMed2000

Identification of age-dependent changes in expression of senescence-accelerated mouse (SAMP8) hippocampal proteins by expression array analysis.

Aging is associated with extensive cognitive impairments, although the biochemical and physiological basis of these deficits are unknown. As the hippocampus plays a vital role in cognitive functions, we have selected this tissue to analyze changes in gene expression at two different ages. Array technology is utilized to explore how gene expression in hippocampus is affected by accelerated cognitive impairment in Senescence-Accelerated Mouse (SAM P8) strain. We show that the expression of genes associated with stress response and xenobiotic metabolism are strongly affected at a time when cognitive impairment occurs. Affected genes include those involved both in signaling and chaperone function. The effector and regulator family of chaperones, which play an important role in protein folding, and also the xenobiotic metabolizing enzymes that play crucial role in antioxidant systems, show significant changes in gene expression between 4 and 12 months.

Aging↗
PubMed2002

High-density microarray analysis of hippocampal gene expression following experimental brain injury.

Behavioral, biophysical, and pharmacological studies have implicated the hippocampus in the formation and storage of spatial memory. Traumatic brain injury (TBI) often causes spatial memory deficits, which are thought to arise from the death as well as the dysfunction of hippocampal neurons. Cell death and dysfunction are commonly associated with and often caused by altered expression of specific genes. The identification of the genes involved in these processes, as well as those participating in postinjury cellular repair and plasticity, is important for the development of mechanism-based therapies. To monitor the expression levels of a large number of genes and to identify genes not previously implicated in TBI pathophysiology, a high-density oligonucleotide array containing 8,800 genes was interrogated. RNA samples were prepared from ipsilateral hippocampi 3 hr and 24 hr following lateral cortical impact injury and compared to samples from sham-operated controls. Cluster analysis was employed using statistical algorithms to arrange the genes according to similarity in patterns of expression. The study indicates that the genomic response to TBI is complex, affecting approximately 6% (at the time points examined) of the total number of genes examined. The identity of the genes revealed that TBI affects many aspects of cell physiology, including oxidative stress, metabolism, inflammation, structural changes, and cellular signaling. The analysis revealed genes whose expression levels have been reported to be altered in response to injury as well as several genes not previously implicated in TBI pathophysiology.

Animals↗
PubMed2002

Expression profiling of microdissected pancreatic adenocarcinomas.

Pancreatic ductal adenocarcinoma is characterized by a paucity of neoplastic cells embedded in a densely desmoplastic stroma. Therefore, laser capture microdissection was performed to obtain homogeneous populations of normal and neoplastic ductal cells. These were subjected to a comparative study of gene expression utilizing human cDNA arrays. A variety of dysregulated genes were identified, comprising cell cycle and growth regulators, invasion regulators, signalling and developmental molecules. In addition to genes already found to be overexpressed in pancreatic cancer, such as TIMP1, MMP7, CD59, rhoC and NDKA, we present evidence to implicate genes which have not previously been reported in this tumour type. These include the overexpressed genes ABL2, Notch4 and SOD1, as well as XRCC1, a DNA repair gene whose transcript was found downregulated. Quantitative real-time RT-PCR (QRT-PCR) was employed to confirm differential expression of ABL2, Notch4 and SOD1 and immunohistochemical analysis was used to verify decreased protein expression of XRCC1 using a custom-built pancreatic tissue array. Combining microarray-derived gene expression profiles of pure pancreatic cell populations, QRT-PCR and pancreas-specific tissue arrays therefore proved to be highly informative in elucidating the molecular pathology of this highly malignant tumour type.

Adenocarcinoma↗
PubMed2003

Gene expression in donor corneal endothelium.

OBJECTIVE: To report gene expression profiles of normal human corneal endothelium with microarray analysis and serial analysis of gene expression (SAGE). METHODS: Corneal endothelium was removed from normal human corneas obtained from eye banks. Total RNA was isolated and SAGE analysis was performed. The same RNA source was used to construct a complementary DNA library that was hybridized to microarrays containing 12 558 transcripts. RESULTS: A total of 9530 SAGE tags were sequenced, representing 4724 unique tags. Microarray analysis identified 542 distinct transcripts. A database of human corneal endothelial gene expression was compiled. Of the SAGE tags, 1720 matched known genes, 478 corresponded to expressed sequence tags, and 2526 had no known match to public databases. The 5 most abundantly expressed SAGE tags were cytochrome c oxidase subunit II, adenosine triphosphate synthase F(0) subunit 6, carbonic anhydrase XII, 12S ribosomal RNA, and ferritin, heavy polypeptide 1. Thirty-four percent of the transcripts (n = 1616) were specific to the corneal endothelium, when compared with other publicly available SAGE libraries. The 5 most abundant unique tags were keratin 12, angiopoietinlike factor, annexin A8, and 2 tags with no match to the database. Many endothelial pump function enzymes were confirmed, including several plasma membrane Na( +)/K(+) adenosine triphosphatases and a recently reported bicarbonate transporter. CONCLUSIONS: Corneal endothelial gene expression profiles by the current analysis provide an understanding of endothelial metabolism, structure, and function; enable comparisons to diseased endothelium; and provide baseline data that may lead to the discovery of novel endothelial genes.

Aged↗
PubMed2001

Anabolic and catabolic gene expression pattern analysis in normal versus osteoarthritic cartilage using complementary DNA-array technology.

OBJECTIVE: To understand changes in gene expression levels that occur during osteoarthritic (OA) cartilage degeneration, using complementary DNA (cDNA)-array technology. METHODS: Nine normal, 6 early degenerated, and 6 late-stage OA cartilage samples of human knee joints were analyzed using the Human Cancer 1.2 cDNA array and TaqMan analysis. RESULTS: In addition to a large variability of expression levels between different patients, significant expression patterns were detectable for many genes. Cartilage types II and VI collagen were strongly expressed in late-stage specimens, reflecting the high matrix-remodeling activity of advanced OA cartilage. The increase in fibronectin expression in early degeneration suggests that fibronectin is a crucial regulator of matrix turnover activity of chondrocytes during early disease development. Of the matrix metalloproteinases (MMPs), MMP-3 appeared to be strongly expressed in normal and early degenerative cartilage and down-regulated in the late stages of disease. This indicates that other degradation pathways might be more important in late stages of cartilage degeneration, involving other enzymes, such as MMP-2 and MMP-11, both of which were up-regulated in late-stage disease. MMP-11 was up-regulated in OA chondrocytes and, interestingly, also in the early-stage samples. Neither MMP-1 nor MMP-8 was detectable, and MMP-13 and MMP-2 were significantly detectable only in late-stage specimens, suggesting that early stages are characterized more by degradation of other matrix components, such as aggrecan and other noncollagenous molecules, than by degradation of type II collagen fibers. CONCLUSION: This investigation allowed us to identify gene expression profiles of the disease process and to get new insights into disease mechanisms, for example, to develop a picture of matrix proteinases that are differentially involved in different phases of the disease process.

Adult↗
PubMed2003

Cell and organ printing 1: protein and cell printers.

We have developed several devices for positioning organic molecules, molecular aggregates, cells, and single-cell organisms onto solid supports. These printers can create stable, functional protein arrays using an inexpensive technology. The cell printer allows us to create cell libraries as well as cellular assemblies that mimic their respective position in organs. The printers are derived from commercially available ink-jet printers that are modified to dispense protein or cell solutions instead of ink. We describe here the modifications to the print heads, and the printer hardware and software that enabled us to adapt the ink-jet printers for the manufacture of cell and protein arrays. The printers have the advantage of being fully automated and computer controlled, and allow for the high-throughput manufacture of protein and cell arrays.

Animals↗
PubMed2002

Neuropilin-1 is differentially expressed in myoepithelial cells and vascular smooth muscle cells in preneoplastic and neoplastic human breast: a possible marker for the progression of breast cancer.

The expression and distribution of neuropilin-1 (NRP-1) was examined in the samples of normal human breast tissues and in non-neoplastic and neoplastic areas of breast tissue removed for carcinoma using RT-PCR as well as conventional and tissue microarrays immunohistochemical analyses. The NRP-1 mRNA expression was significantly higher in neoplastic tissues as compared to normal breast samples. Immunohistochemically, the myoepithelial cells of the mammary ducts and lobules display positive reactions for NRP-1, whereas the inner ductal and lobular epithelial cell layers failed to react. The myoepithelial cells of ducts and lobules in both neoplastic and non-neoplastic tissue specimens displayed a stronger positive reaction for NRP-1 than those in the normal breast. A positive reaction for NRP-1, but with a gradual reduction in intensity, was observed in the myoepithelial cells of ducts with atypical epithelial hyperplasia and ductal carcinoma in situ (DCIS). The reaction was undetected or minimally detected in the areas of invasive carcinoma. NRP-1 positive immunolabeling was also localized in the vascular smooth muscle cells and in some endothelial cells of the blood vessels in normal, non-neoplastic and neoplastic breast tissue samples. In areas of breast carcinoma, NRP-1 immunolabeling was more prominent in both vascular smooth muscle cells and in some endothelial cells than in similar cells in normal breast. The specificity of the newly developed antibody for NRP-1 was confirmed by in situ hybridization with DIG-labeled PCR generated probe. These results suggest that NRP-1 may be a multiple function protein in human breast and may be involved in the induction of local invasiveness of neoplasia and angiogenesis and have direct relevance to the progression of breast cancer.

Base Sequence↗
PubMed2001

Gene expression profiling in postmortem Rett Syndrome brain: differential gene expression and patient classification.

The identification of mutations in the transcriptional repressor methyl-CpG-binding protein 2 (MECP2) gene in Rett Syndrome (RTT) suggests that an inappropriate release of transcriptional silencing may give rise to RTT neuropathology. Despite this progress, the molecular basis of RTT neuropathogenesis remains unclear. Using multiple cDNA microarray technologies, subtractive hybridization, and conventional biochemistry, we generated comprehensive gene expression profiles of postmortem brain tissue from RTT patients and matched controls. Many glial transcripts involved in known neuropathological mechanisms were found to have increased expression in RTT brain, while decreases were observed in the expression of multiple neuron-specific mRNAs. Dramatic and consistent decreases in transcripts encoding presynaptic markers indicated a specific deficit in presynaptic development. Employing multiple clustering algorithms, it was possible to accurately segregate RTT from control brain tissue samples based solely on gene expression profile. Although previously achieved in cancers, our results constitute the first report of human disease classification using gene expression profiling in a complex tissue source such as brain.

Adolescent↗
PubMed2003

Genetic animal models of anxiety.

The focus of this review is on progress achieved in identifying specific genes conferring risk for anxiety disorders through the use of genetic animal models. We discuss gene-finding studies as well as those manipulating a candidate gene. Both human and animal studies thus far support the genetic complexity of anxiety. Clinical manifestations of these diseases are likely related to multiple genes. While different anxiety disorders and anxiety-related traits all appear to be genetically influenced, it has been difficult to ascertain genetic influences in common. Mouse studies have provisionally mapped several loci harboring genes that affect anxiety-related behavior. The growing array of mutant mice is providing valuable information about how genes and environment interact to affect anxious behavior via multiple neuropharmacological pathways. Classical genetic methods such as artificial selection of rodents for high or low anxiety are being employed. Expression array technologies have as yet not been employed, but can be expected to implicate novel candidates and neurobiological pathways.

Animals↗