Statin-induced changes in gene expression in EBV-transformed and native B-cells.

Human lymphoblastoid cell lines (LCLs), generated through Epstein-Barr Virus (EBV) transformation of B-lymphocytes (B-cells), are a commonly used model system for identifying genetic influences on human diseases and on drug responses. We have previously used LCLs to examine the cellular effects of genetic variants that modulate the efficacy of statins, the most prescribed class of cholesterol-lowering drugs used for the prevention and treatment of cardiovascular disease. However, statin-induced gene expression differences observed in LCLs may be influenced by their transformation, and thus differ from those observed in native B-cells. To assess this possibility, we prepared LCLs and purified B-cells from the same donors, and compared mRNA profiles after 24 h incubation with simvastatin (2 µm) or sham buffer. Genes involved in cholesterol metabolism were similarly regulated between the two cell types under both the statin and sham-treated conditions, and the statin-induced changes were significantly correlated. Genes whose expression differed between the native and transformed cells were primarily implicated in cell cycle, apoptosis and alternative splicing. We found that ChIP-seq signals for MYC and EBNA2 (an EBV transcriptional co-activator) were significantly enriched in the promoters of genes up-regulated in the LCLs compared with the B-cells, and could be involved in the regulation of cell cycle and alternative splicing. Taken together, the results support the use of LCLs for the study of statin effects on cholesterol metabolism, but suggest that drug effects on cell cycle, apoptosis and alternative splicing may be affected by EBV transformation. This dataset is now uploaded to GEO at the accession number GSE51444.

Hepatic nuclear factor 1-alpha: inflammation, genetics, and atherosclerosis.

PURPOSE OF REVIEW: Increased plasma levels of C-reactive protein (CRP), a hepatic acute phase reactant, predict risk for coronary heart disease. There has been interest in identifying genetic determinants of CRP as a means of better understanding its regulation and its relation to coronary heart disease. We here review recent findings that have linked plasma CRP levels to single nucleotide polymorphisms in hepatic nuclear factor (HNF) 1-alpha, a transcription factor with a wide range of functions, including many involved in cholesterol, bile acid, and lipoprotein metabolism. RECENT FINDINGS: Two genome-wide association studies have identified single nucleotide polymorphisms in several genes that are strongly related to plasma CRP levels, including several on chromosome 12 in the vicinity of the HNF1A gene. The CRP gene promoter has two HNF1-alpha-binding sites. Recently, it has been demonstrated that HNF1-alpha is required for cytokine-driven CRP expression and that this involves formation of a complex with STAT3 and c-Fos. SUMMARY: Based on the recent genetic findings as well as delineation of the role of HNF1-alpha in regulating the expression of the CRP gene, it appears that this transcription factor may play a key role in linking metabolic and inflammatory pathways underlying the pathogenesis of coronary heart disease.

Developing Interactive Exhibits with Scientists: Three Example Collaborations from the Life Sciences Collection at the Exploratorium.

Science museums have made a concerted effort to work with researchers to incorporate current scientific findings and practices into informal learning opportunities for museum visitors. Many of these efforts have focused on creating opportunities and support for researchers to interact face-to-face with the public through, for example, speaker series, community forums, and engineering competitions. However, there are other means by which practicing scientists can find a voice on the museum floor-through the design and development of exhibits. Here we describe how researchers and museum professionals have worked together to create innovative exhibit experiences for an interactive science museum. For each example: scientist as (1) data providers, (2) advisors, and (3) co-developers, we highlight essential components for a successful partnership and pitfalls to avoid when collaborating on museum exhibits. Not many museums prototype and build their own exhibits like the Exploratorium. In those cases, there may be similar opportunities in more mediated offerings such as public demonstrations or lectures or in other formats that allow for direct interactions between scientists and visitors. We believe there are many opportunities for researchers to share natural phenomena, to advise on exhibit development and interpretation, to provide much needed materials, and to otherwise incorporate authentic research into the learning experiences at museums, no matter what the format.

Gene expression profiling in chronic copper overload reveals upregulation of Prnp and App.

The level at which copper becomes toxic is not clear. Several studies have indicated that copper causes oxidative stress; however, most have tested very high levels of copper exposure. We currently have only a limited understanding of the protective systems that operate in cells chronically exposed to copper. Additionally, the limits of homeostatic regulation are not known, making it difficult to define the milder effects of copper excess. Furthermore, a robust assay to facilitate the diagnosis of copper excess and to distinguish mild, moderate, and severe copper overload is needed. To address these issues, we have investigated the effects on steady-state gene expression of chronic copper overload in a cell culture model system using cDNA microarrays. For this study we utilized cells from genetic models of copper overload: fibroblast cells from two mouse mutants, C57BL/6-Atp7a(Mobr) and C57BL/6-Atp7a(Modap). These cell lines accumulate copper to abnormally high levels in normal culture media due to a defect in copper export from the cell. We identified 12 differentially expressed genes in common using our outlier identification methods. Surprisingly, our results show no evidence of oxidative stress in the copper-loaded cells. In addition, candidate components perhaps responsible for a copper-specific homeostatic response are identified. The genes that encode for the prion protein and the amyloid-beta precursor protein, two known copper-binding proteins, are upregulated in both cell lines.

Gene expression profiling in wild-type and metallothionein mutant fibroblast cell lines.

The role of metallothioneins (MT) in copper homeostasis is of great interest, as it appears to be partially responsible for the regulation of intracellular copper levels during adaptation to extracellular excess of the metal. To further investigate a possible role of MTs in copper metabolism, a genomics approach was utilized to evaluate the role of MT on gene expression. Microarray analysis was used to examine the effects of copper overload in fibroblast cells from normal and MT I and II double knock-out mice (MT-/-). As a first step, we compared genes that were significantly upregulated in wild-type and MT-/- cells exposed to copper. Even though wild-type and mutant cells are undistinguishable in terms of their morphological features and rates of growth, our results show that MT-/- cells do not respond with induction of typical markers of cellular stress under copper excess conditions, as observed in the wild-type cell line, suggesting that the transcription initiation rate or the mRNA stability of stress genes is affected when there is an alteration in the copper store capacity. The functional classification of other up-regulated genes in both cell lines indicates that a large proportion (>80%) belong to two major categories: 1) metabolism; and 2) cellular physiological processes, suggesting that at the transcriptional level copper overload induces the expression of genes associated with diverse molecular functions. These results open the possibility to understand how copper homeostasis is being coordinated with other metabolic pathways.

Gene expression profiling in wild-type and metallothionein mutant fibroblast cell lines

The role of metallothioneins (MT) in copper homeostasis is of great interest, as it appears to be partially responsible for the regulation of intracellular copper levels during adaptation to extracellular excess of the metal. To further investigate a possible role of MTs in copper metabolism, a genomics approach was utilized to evaluate the role of MT on gene expression. Microarray analysis was used to examine the effects of copper overload in fibroblast cells from normal and MT I and II double knock-out mice (MT-/-). As a first step, we compared genes that were significantly upregulated in wild-type and MT-/- cells exposed to copper. Even though wild-type and mutant cells are undistinguishable in terms of their morphological features and rates of growth, our results show that MT-/- cells do not respond with induction of typical markers of cellular stress under copper excess conditions, as observed in the wild-type cell line, suggesting that the transcription initiation rate or the mRNA stability of stress genes is affected when there is an alteration in the copper store capacity. The functional classification of other up-regulated genes in both cell lines indicates that a large proportion (>80 percent) belong to two major categories: 1) metabolism; and 2) cellular physiological processes, suggesting that at the transcriptional level copper overload induces the expression of genes associated with diverse molecular functions. These results open the possibility to understand how copper homeostasis is being coordinated with other metabolic pathways.