Project description:Precision mass spectrometry, animal models and organ-on-a-chip (OOC) systems have emerged as promising experimental strategies to study the molecular mechanisms governing human biology and disease, including the molecular basis of cardiovascular disorders like fibrosis where access to patient samples is limiting and subject to confounding variables. Nevertheless, no systematic comparisons have ever been reported, precluding an objective assessment of cross-platform consistency, performance and bias. Here, we apply and evaluate an integrative mass spectrometry-based platform that allows for quantitative global phospho/proteomic surveys of normal and afflicted tissue from human, mouse and OOC-derived specimens. The applicability and utility of this approach was tested in the context of cardiac fibrosis through comprehensive analyses of fibrotic cardiomyocyte samples from Biowire OOC specimens, and cardiac tissue explants from hypertrophic patients and a mouse genetic model, and show that clinically meaningful biological inferences can be generated by leveraging commonalities and unique attributes generated across each platform.
Project description:Pre-clinical models that effectively recapitulate human disease are critical for expanding our knowledge of cancer biology and drug resistance mechanisms. For haematological malignancies, the non-obese diabetic/severe combined immunodeficient (NOD/SCID) mouse is one of the most successful models to study paediatric acute lymphoblastic leukaemia (ALL). However, for this model to be effective for studying engraftment and therapy responses at the whole genome level, careful molecular characterisation is essential.Here, we sought to validate species-specific gene expression profiling in the high engraftment continuous ALL NOD/SCID xenograft. Using the human Affymetrix whole transcript platform we analysed transcriptional profiles from engrafted tissues without prior cell separation of mouse cells and found it to return highly reproducible profiles in xenografts from individual mice. The model was further tested with experimental mixtures of human and mouse cells, demonstrating that the presence of mouse cells does not significantly skew expression profiles when xenografts contain 90% or more human cells. In addition, we present a novel in silico and experimental masking approach to identify probes and transcript clusters susceptible to cross-species hybridisation.We demonstrate species-specific transcriptional profiles can be obtained from xenografts when high levels of engraftment are achieved or with the application of transcript cluster masks. Importantly, this masking approach can be applied and adapted to other xenograft models where human tissue infiltration is lower. This model provides a powerful platform for identifying genes and pathways associated with ALL disease progression and response to therapy in vivo.
Project description:Microarray technology has had a profound impact on gene expression research. Some studies have questioned whether similar expression results are obtained when the same RNA samples are analyzed on different platforms. The MicroArray Quality Control (MAQC) project was initiated to address these concerns, as well as other performance and analysis issues. We demonstrate the consistency of results within a platform across test sites as well as the high level of cross-platform concordance in terms of genes identified as differentially expressed. The MAQC study provides a rich resource that will help build consensus on the use of microarrays in research, clinical and regulatory settings. Manuscripts related to the MAQC project have been published in Nature Biotechnology, 24(9), September, 2006. More information about the MAQC project can be found at http://edkb.fda.gov/MAQC/. Keywords: Cross-platform comparison
Project description:Leveraging the conserved cancer genomes across mammals has the potential to transform driver gene discovery in orphan cancers. Here, we combine cross-species genomics with validation across human-dog-mouse systems to uncover a new osteosarcoma driver genes. This widely applicable cross-species approach serves as a platform to expedite search of cancer drivers in rare human malignancies offering new targets for cancer therapy.
Project description:Mesenchymal stem cells (MSC) are the most commonly used cells in tissue engineering and regenerative medicine. MSC can promote host tissue repair through several different mechanisms including donor cell engraftment, release of cell signaling factors and the transfer of healthy organelles to the host. In the present study, we examine the specific impacts of MSC on mitochondrial morphology and function in host tissues. Employing in vitro cell culture of inherited mitochondrial disease and an in vivo animal experimental model of low-grade inflammation (high fat feeding), we show human-derived MSC to alter mitochondrial function. MSC co-culture of skin fibroblasts from mitochondrial disease patients rescued aberrant mitochondrial morphology from a fission state to a more fused appearance indicating an effect of MSC co-culture on host cell mitochondrial network formation. In vivo experiments confirmed mitochondrial abundance and mitochondrial oxygen consumption rates were elevated in host tissues following MSC treatment. Furthermore, microarray profiling identified 226 genes with differential expression in the liver of animals treated with MSC, with cellular signaling and actin cytoskeleton regulation as key upregulated processes. Collectively, our data indicate that MSC therapy rescues impaired mitochondrial morphology, enhances host metabolic capacity and induces widespread host gene shifting. These results highlight the potential of MSC to modulate mitochondria in both inherited and pathological disease states. To examine the effect of MSC therapy on global gene expression and cell signaling pathways, microarray gene expression analysis was performed on liver tissues.
Project description:The comparative whole genome transcriptome effects of two similar pharmaceuticals, imig or vela, on a Gaucher disease mouse model, 9V/null, were evaluated by two commonly used platforms, mRNA-Seq and microarray. Also, statistical methods, DESeq and edgeR for mRNA-Seq and Mixed Model ANOVA for microarray, were compared for differential gene expression detection. The biological pathways were similar between two platforms. Cell growth and proliferation, cell cycle, heme metabolism, and mitochondrial dysfunction were the most altered functions associated with the disease process. Although the two biopharmaceuticals have a very similar structure and function, imig- and vela- treatment in the mice differentially affected disease-specific pathways indicating the action of the two drugs on the disease process in the visceral tissues of Gaucher mouse model differ significantly at the molecular level. This study provides a comprehensive comparison between the two platforms (mRNA-Seq and microarray) for gene expression analysis and addresses the contribution of the different methods involved in the analysis of such data. The results also provide insights into the differential molecular effects of two similar biopharmaceuticals for Gaucher disease treatment 9V/null mice (Gaucher mouse model) were injected weekly via tail vein with 60U/kg/wk of imig or vela for 8 wks. Control 9V/null mice were injected with same volume of saline. Wt mice were untreated. Age and strain matched mice were used for the study. Also, statistical methods, DESeq and edgeR for mRNA-Seq and Mixed Model ANOVA for microarray, were compared for differential gene expression detection. Cell growth and proliferation, cell cycle, heme metabolism, and mitochondrial dysfunction were the most altered functions associated with the disease process. The results also provide insights into the differential molecular effects of two similar biopharmaceuticals for Gaucher disease treatment.