Gene expression profiling of cardiovascular disease models

Recent development of gene expression profiling technologies has enabled the large-scale analysis of gene expression changes during disease progression. Frequently, cardiovascular diseases involve complex interactions of multiple cell types over prolonged periods of time. A better understanding of the pathology of cardiovascular diseases and the potential identification of underlying genetic defects are currently being explored by using profiling methodologies in a number of animal and tissue-culture models.     

Gene expression profiling in autoimmune noninfectious uveitis disease

Noninfectious uveitis is a predominantly T cell-mediated autoimmune, intraocular inflammatory disease. To characterize the gene expression profile from patients with noninfectious uveitis, PBMCs were isolated from 50 patients with clinically characterized noninfectious uveitis syndrome. A pathway-specific cDNA microarray was used for gene expression profiling and real-time PCR array for further confirmation. Sixty-seven inflammation- and autoimmune-associated genes were found differentially expressed in uveitis patients, with 28 of those genes being validated by real-time PCR. Several genes previously unknown for autoimmune uveitis, including IL-22, IL-19, IL-20, and IL-25/IL-17E, were found to be highly expressed among uveitis patients compared with the normal subjects with IL-22 expression highly variable among the patients. Furthermore, we show that IL-22 can affect primary human retinal pigment epithelial cells by decreasing total tissue resistance and inducing apoptosis possibly by decreasing phospho-Bad level. In addition, the microarray data identified a possible uveitis-associated gene expression pattern, showed distinct gene expression profiles in patients during periods of clinical activity and quiescence, and demonstrated similar expression patterns in related patients with similar clinical phenotypes. Our data provide the first evidence that a subset of IL-10 family genes are implicated in noninfectious uveitis and that IL-22 can affect human retinal pigment epithelial cells. The results may facilitate further understanding of the molecular mechanisms of autoimmune uveitis and other autoimmune originated inflammatory diseases.     

Gene expression profiling of rewarding effect in methamphetamine treated Bax-deficient mouse

Methamphetamine is an illicit drug that is often abused and can cause neuropsychiatric and neurotoxic damage. Repeated administration of psychostimulants such as methamphetamine induces a behavioral sensitization. According to a previous study, Bax was involved in neurotoxicity by methamphetamine, but the function of Bax in rewarding effect has not yet been elucidated. Therefore, we have studied the function of Bax in a rewarding effect model. In the present study, we treated chronic methamphetamine exposure in a Bax-deficient mouse model and examined behavioral change using a conditioned place preference (CPP) test. The CPP score in Bax knockout mice was decreased compared to that of wild-type mice. Therefore, we screened for Bax-related genes that are involved in rewarding effect using microarray technology. In order to confirm microarray data, we applied the RT-PCR method to observe relative changes of Bcl2, a pro-apoptotic family gene. As a result, using our experiment microarray, we selected genes that were associated with Bax in microarray data, and eventually selected the Tgfbr2 gene. Expression of the Tgfbr2 gene was decreased by methamphetamine in Bax knockout mice, and the gene was overexpressed in Bax wild-type mice. Additionally, we confirmed that Creb, FosB, and c-Fos were related to rewarding effect and Bax using immunohistochemistry.     

Gene expression profiling of coronary artery disease and its relation with different severities

Global gene expression profiling is a powerful tool enabling the understanding of pathophysiology and subsequent management of diseases. This study aims to explore functionally annotated differentially expressed genes (DEGs); their biological processes for coronary artery disease (CAD) and its different severities of atherosclerotic lesions. This study also aims to identify the change in expression patterns of DEGs in atherosclerotic lesions of single-vessel disease (SVD) and triple-vessel disease (TVD). The weight of different severities of lesion was estimated using a modified Gensini score. The gene expression profiling was performed using the Affymetrix microarray platform. The functional annotation for CAD was performed using DAVID v6.8. The biological network gene ontology tool (BiNGO) and ClueGO were used to explore the biological processes of functionally annotated genes of CAD. The changes in gene expression from SVD to TVD were determined by evaluating the fold change. Functionally annotated genes were found in an unique set and could be distinguishing two distinct severities of CAD. The biological processes such as cellular migration, locomotion, cell adhesion, cytokine production, positive regulation of cell death etc. enriched the functionally annotated genes in SVD, whereas, wound healing, negative regulation of cell death, blood coagulation, angiogenesis and fibrinolysis were enriched significantly in TVD patients. The genes THBS1 and CAPN10 were functionally annotated for CAD in both SVD and TVD. The 61 DEGs were identified, those have changes their expression with different severities of atherosclerotic lesions, in which 13 genes had more than two-fold change in expression between SVD and TVD. The consistent findings were obtained on validation of microarray gene expression of selected 10 genes in a separate cohort using real-time PCR. This study identified putative candidate genes and their biological processes predisposing toward and affecting the severity of CAD.     

Gene expression profiling of the rewarding effect caused by methamphetamine in the mesolimbic dopamine system

Methamphetamine, a commonly used addictive drug, is a powerful addictive stimulant that dramatically affects the CNS. Repeated METH administration leads to a rewarding effect in a state of addiction that includes sensitization, dependence, and other phenomena. It is well known that susceptibility to the development of addiction is influenced by sources of reinforcement, variable neuroadaptive mechanisms, and neurochemical changes that together lead to altered homeostasis of the brain reward system. These behavioral abnormalities reflect neuroadaptive changes in signal transduction function and cellular gene expression produced by repeated drug exposure. To provide a better understanding of addiction and the mechanism of the rewarding effect, it is important to identify related genes. In the present study, we performed gene expression profiling using microarray analysis in a reward effect animal model. We also investigated gene expression in four important regions of the brain, the nucleus accumbens, striatum, hippocampus, and cingulated cortex, and analyzed the data by two clustering methods. Genes related to signaling pathways including G-protein-coupled receptor-related pathways predominated among the identified genes. The genes identified in our study may contribute to the development of a gene modeling network for methamphetamine addiction.     

Gene expression profiling in porcine fetal thymus

To obtain an initial overview of gene diversity and expression pattern in porcine thymus, 11,712 ESTs (Expressed Sequence Tags) from 100-day-old porcine thymus (FTY) were sequenced and 7,071 cleaned ESTs were used for gene expression analysis. Clustered by the PHRAP program, 959 contigs and 3,074 singlets were obtained. Blast search showed that 806 contigs and 1,669 singlets (totally 5,442 ESTs) had homologues in GenBank and 1,629 ESTs were novel. According to the Gene Ontology classification, 36.99% ESTs were cataloged into the gene expression group, indicating that although the functional gene (18.78% in defense group) of thymus is expressed in a certain degree, the 100-day-old porcine thymus still exists in a developmental stage. Comparative analysis showed that the gene expression pattern of the 100-day-old porcine thymus is similar to that of the human infant thymus.     

毒品成瘾与脑组织基因表达谱的研究进展

毒品成瘾是由滥用毒品外在因素与遗传易感性等内在因素共同作用而导致的一种慢性脑疾病; 毒品成瘾的机制目前还不十分清楚。毒品成瘾研究的一个主要目标是鉴别和分离毒品导致脑功能障碍的分子机制; 采用高通量的基因表达谱技术研究毒品成瘾者在不同状态下脑基因表达全貌, 对深入认识毒品成瘾的机制具有重要的意义。文章综述了毒品成瘾的遗传机制及高通量脑组织基因组表达技术——SAGE和微阵列(Microarry)在毒品成瘾研究中的进展。     

Gene expression profiling of the preclinical scrapie-infected hippocampus

The molecular events that underlie prion disease neuropathology remain poorly defined. Within the hippocampus of the ME7/CV mouse scrapie model, profound CA1 neuronal loss occurs between 160 and 180 days post-infection (dpi). To elucidate the molecular events that may contribute to this neuronal loss, we have applied Affymetrix high-density oligonucleotide probe arrays to the study of ME7-infected hippocampal gene expression at 170 dpi. The study has identified 78 genes that are differentially expressed greater than 1.5-fold within the preclinical ME7-infected hippocampus prior to the profound late stage glial cell activation. The results indicate oxidative and endoplasmic reticulum (ER) stress, activated ER and mitochondrial apoptosis pathways, and activated cholesterol biosynthesis within the scrapie-infected hippocampus, and offer insight into the molecular events which underlie the neuropathology.     

Gene expression profiling in the inductive human hematopoietic microenvironment

Human hematopoietic stem cells (HSCs) and their progenitors can be maintained in vitro in long-term bone marrow cultures (LTBMCs) in which constituent HSCs can persist within the adherent layers for up to 2 months. Media replenishment of LTBMCs has been shown to induce transition of HSCs from a quiescent state to an active cycling state. We hypothesize that the media replenishment of the LTBMCs leads to the activation of important regulatory genes uniquely involved in HSC proliferation and differentiation. To profile the gene expression changes associated with HSC activation, we performed suppression subtractive hybridization (SSH) on day 14 human LTBMCs following 1-h media replenishment and on unmanipulated controls. The generated SSH library contained 191 differentially up-regulated expressed sequence tags (ESTs), the majority corresponding to known genes related to various intracellular processes, including signal transduction pathways, protein synthesis, and cell cycle regulation. Nineteen ESTs represented previously undescribed sequences encoding proteins of unknown function. Differential up-regulation of representative genes, including IL-8, IL-1, putative cytokine 21/HC21, MAD3, and a novel EST was confirmed by semi-quantitative RT-PCR. Levels of fibronectin, G-CSF, and stem cell factor also increased in the conditioned media of LTBMCs as assessed by ELISA, indicating increased synthesis and secretion of these factors. Analysis of our library provides insights into some of the immediate early gene changes underlying the mechanisms by which the stromal elements within the LTBMCs contribute to the induction of HSC activation and provides the opportunity to identify as yet unrecognized factors regulating HSC activation in the LTBMC milieu.     

Gene expression profiling of hereditary exencephaly in chickens

In this preliminary study, differentially expressed genes were investigated in cranial tissues from chickens with hereditary exencephaly using cDNA microarrays containing 1,152 genes and expressed sequence tags (ESTs). Genes showing twofold or greater differences at P < 0.05 between affected and normal cranial cells were considered to be candidates for hereditary exencephaly in chicken. Eighteen ESTs (11 known genes/homologues) were upregulated and 108 ESTs (51 known genes/homologues) were downregulated. The EST AL584231 (ROS006C9), orthologous to human MTHFD1, a known candidate gene for human neural tube defects (NTDs), was expressed at the same level both in normal and affected chicken cranial tissues. ESTs AL584253 (ROS006F7, thioredoxin reductase 1) and AL585511 (ROS024H9, thioredoxin), both involved in NTD pathogenic pathways in mice, were downregulated and had mean ratios of 0.41 and 0.04 for expression in affected vs. normal cells respectively. Expression differences of these two ESTs were confirmed by quantitative real-time polymerase chain reaction. These data indicate that ESTs AL584253 and AL585511 are candidates for hereditary exencephaly in chickens.     

Gene expression profiling in mitochondrial disease: assessment of microarray accuracy by high-throughput Q-PCR

Mitochondrial diseases are a heterogeneous array of disorders with a complex etiology. Use of microarrays as a tool to investigate complex human disease is increasingly common, however, a principle drawback of microarrays is their limited dynamic range, due to the poor quantification of weak signals. Although it is generally understood that low-intensity microarray 'spots' may be unreliable, there exists little documentation of their accuracy. Quantitative PCR (Q-PCR) is frequently used to validate microarray data, yet few Q-PCR validation studies have focused on the accuracy of low-intensity microarray signals. Hence, we have used Q-PCR to systematically assess microarray accuracy as a function of signal strength in a mouse model of mitochondrial disease, the superoxide dismutase 2 (SOD2) nullizygous mouse. We have focused on a unique category of data--spots with only one weak signal in a two-dye comparative hybridization--and show that such 'high-low' signal intensities are common for differentially expressed genes. This category of differential expression may be more important in mitochondrial disease in which there are often mosaic expression patterns due to the idiosyncratic distribution of mutant mtDNA in heteroplasmic individuals. Using RNA from the SOD2 mouse, we found that when spotted cDNA microarray data are filtered for quality (low variance between many technical replicates) and spot intensity (above a negative control threshold in both channels), there is an excellent quantitative concordance with Q-PCR (R2 = 0.94). The accuracy of gene expression ratios from low-intensity spots (R2 = 0.27) and 'high-low' spots (R2 = 0.32) is considerably lower. Our results should serve as guidelines for microarray interpretation and the selection of genes for validation in mitochondrial disorders.     

Gene expression profiling of the pH response in Escherichia coli

Escherichia coli MG1655 acid-inducible genes were identified by whole-genome expression profiling. Cultures were grown to the mid-logarithmic phase on acidified glucose minimal medium, conditions that induce glutamate-dependent acid resistance (AR), while the other AR systems are either repressed or not induced. A total of 28 genes were induced in at least two of three experiments in which the gene expression profiles of cells grown in acid (pH 5.5 or 4.5) were compared to those of cells grown at pH 7.4. As expected, the genes encoding glutamate decarboxylase, gadA and gadB, were significantly induced. Interestingly, two acid-inducible genes code for small basic proteins with pIs of >10.5, and six code for small acidic proteins with pIs ranging from 5.7 to 4.0; the roles of these small basic and acidic proteins in acid resistance are unknown. The acid-induced genes represented only five functional grouping categories, including eight genes involved in metabolism, nine associated with cell envelope structures or modifications, two encoding chaperones, six regulatory genes, and six unknown genes. It is unlikely that all of these genes are involved in the glutamate-dependent AR. However, nine acid-inducible genes are clustered in the gadA region, including hdeA, which encodes a putative periplasmic chaperone, and four putative regulatory genes. One of these putative regulators, yhiE, was shown to significantly increase acid resistance when overexpressed in cells that had not been preinduced by growth at pH 5.5, and mutation of yhiE decreased acid resistance; yhiE could therefore encode an activator of AR genes. Thus, the acid-inducible genes clustered in the gadA region appear to be involved in glutatmate-dependent acid resistance, although their specific roles remain to be elucidated.     

Gene expression profiling of scrapie-infected brain tissue

The underlying pathomechanisms in prion infections of the central nervous system are still insufficiently understood. The identification of genes with altered expression patterns in the diseased brain may provide insight into the disease development on the molecular level, which ultimately leads to neuronal loss. To provide a detailed analysis of changes in the molecular level in prion disease pathology we used a large-scale gene array based approach, which covers more than 11,000 functionally characterised sequences and expressed sequence tags, for the analysis of gene expression profile alterations in the cortex, medulla, and pons of scrapie-infected mice. The study identified in total 114 genes with altered mRNA levels, the majority of which were previously not known to be affected by the disease. Overall the gene array data demonstrate the presence of a strong inflammatory reaction and stress response, and show similarities to gene expression patterns found in brains affected by Alzheimer's disease and aging, respectively.