Comparative analysis and integrative classification of NCI60 cell lines and primary tumors using gene expression profiling data

BACKGROUND: NCI60 cell lines are derived from cancers of 9 tissue origins and have been invaluable in vitro models for cancer research and anti-cancer drug screen. Although extensive studies have been carried out to assess the molecular features of NCI60 cell lines related to cancer and their sensitivities to more than 100,000 chemical compounds, it remains unclear if and how well these cell lines represent or model their tumor tissues of origin. Identification and confirmation of correct origins of NCI60 cell lines are critical to their usage as model systems and to translate in vitro studies into clinical potentials. Here we report a direct comparison between NCI60 cell lines and primary tumors by analyzing global gene expression profiles. RESULTS: Comparative analysis suggested that 51 of 59 cell lines we analyzed represent their presumed tumors of origin. Taking advantage of available clinical information of primary tumor samples used to generate gene expression profiling data, we further classified those cell lines with the correct origins into different subtypes of cancer or different stages in cancer development. For example, 6 of 7 non-small cell lung cancer cell lines were classified as lung adenocarcinomas and all of them were classified into late stages in tumor progression. CONCLUSION: Taken together, we developed and applied a novel approach for systematic comparative analysis and integrative classification of NCI60 cell lines and primary tumors. Our results could provide guidance to the selection of appropriate cell lines for cancer research and pharmaceutical compound screenings. Moreover, this gene expression profile based approach can be generally applied to evaluate experimental model systems such as cell lines and animal models for human diseases

ELISA for Aging Biomarkers Induced by Telomere Dysfunction in Human Plasma

Background. We identified cathelicidin related antimicrobial protein (CRAMP) secreted from telomere dysfunctional bone marrow cells of late generation telomerase knockout mice (G4mTerc−/−), increased in blood and various tissues. It can represented human aging and disease. The main aim of this study is to investigate the sensitive direct enzyme-linked immunosorbent assay (ELISA) method to analyze the human aging and disease in plasma and the detailed methods to quantify the direct ELISA of these aging biomarkers. Methods. Telomere lengths of 50 healthy persons are measured with real-time PCR in blood cells. Plasma samples from all subjects are analyzed using direct ELISA. Results. From 25 years old person to 78 years, the telomere length becomes shorter during aging. In blood plasma, the expression levels of CRAMP increases during human aging. There is the reverse correspondence between the telomere length and the plasma CRAMP level. We also find that the fresh plasma, the frozen plasma which thawed less than 3 times, and the plasma kept in the room temperature less than 3 hours are better for the ELISA analyze of CRAMP in the plasma. Conclusion. This CRAMP ELISA could become a powerful tool for investigating the relationship between human aging and telomere length shortening

Developing and applying a gene functional association network for anti-angiogenic kinase inhibitor activity assessment in an angiogenesis co-culture model

<p>Abstract</p> <p>Background</p> <p>Tumor angiogenesis is a highly regulated process involving intercellular communication as well as the interactions of multiple downstream signal transduction pathways. Disrupting one or even a few angiogenesis pathways is often insufficient to achieve sustained therapeutic benefits due to the complexity of angiogenesis. Targeting multiple angiogenic pathways has been increasingly recognized as a viable strategy. However, translation of the polypharmacology of a given compound to its antiangiogenic efficacy remains a major technical challenge. Developing a global functional association network among angiogenesis-related genes is much needed to facilitate holistic understanding of angiogenesis and to aid the development of more effective anti-angiogenesis therapeutics.</p> <p>Results</p> <p>We constructed a comprehensive gene functional association network or interactome by transcript profiling an in vitro angiogenesis model, in which human umbilical vein endothelial cells (HUVECs) formed capillary structures when co-cultured with normal human dermal fibroblasts (NHDFs). HUVEC competence and NHDF supportiveness of cord formation were found to be highly cell-passage dependent. An enrichment test of Biological Processes (BP) of differentially expressed genes (DEG) revealed that angiogenesis related BP categories significantly changed with cell passages. Built upon 2012 DEGs identified from two microarray studies, the resulting interactome captured 17226 functional gene associations and displayed characteristics of a scale-free network. The interactome includes the involvement of oncogenes and tumor suppressor genes in angiogenesis. We developed a network walking algorithm to extract connectivity information from the interactome and applied it to simulate the level of network perturbation by three multi-targeted anti-angiogenic kinase inhibitors. Simulated network perturbation correlated with observed anti-angiogenesis activity in a cord formation bioassay.</p> <p>Conclusion</p> <p>We established a comprehensive gene functional association network to model in vitro angiogenesis regulation. The present study provided a proof-of-concept pilot of applying network perturbation analysis to drug phenotypic activity assessment.</p

Identification of blood biomarkers of rheumatoid arthritis by transcript profiling of peripheral blood mononuclear cells from the rat collagen-induced arthritis model

Rheumatoid arthritis (RA) is a chronic debilitating autoimmune disease that results in joint destruction and subsequent loss of function. To better understand its pathogenesis and to facilitate the search for novel RA therapeutics, we profiled the rat model of collagen-induced arthritis (CIA) to discover and characterize blood biomarkers for RA. Peripheral blood mononuclear cells (PBMCs) were purified using a Ficoll gradient at various time points after type II collagen immunization for RNA preparation. Total RNA was processed for a microarray analysis using Affymetrix GeneChip technology. Statistical comparison analyses identified differentially expressed genes that distinguished CIA from control rats. Clustering analyses indicated that gene expression patterns correlated with laboratory indices of disease progression. A set of 28 probe sets showed significant differences in expression between blood from arthritic rats and that from controls at the earliest time after induction, and the difference persisted for the entire time course. Gene Ontology comparison of the present study with previous published murine microarray studies showed conserved Biological Processes during disease induction between the local joint and PBMC responses. Genes known to be involved in autoimmune response and arthritis, such as those encoding Galectin-3, Versican, and Socs3, were identified and validated by quantitative TaqMan RT-PCR analysis using independent blood samples. Finally, immunoblot analysis confirmed that Galectin-3 was secreted over time in plasma as well as in supernatant of cultured tissue synoviocytes of the arthritic rats, which is consistent with disease progression. Our data indicate that gene expression in PBMCs from the CIA model can be utilized to identify candidate blood biomarkers for RA

DNA microarray data integration by ortholog gene analysis reveals potential molecular mechanisms of estrogen-dependent growth of human uterine fibroids

BACKGROUND: Uterine fibroids or leiomyoma are a common benign smooth muscle tumor. The tumor growth is well known to be estrogen-dependent. However, the molecular mechanisms of its estrogen-dependency is not well understood. METHODS: Differentially expressed genes in human uterine fibroids were either retrieved from published papers or from our own statistical analysis of downloaded array data. Probes for the same genes on different Affymetrix chips were mapped based on probe comparison information provided by Affymetrix. Genes identified by two or three array studies were submitted for ortholog analysis. Human and rat ortholog genes were identified by using ortholog gene databases, HomoloGene and TOGA and were confirmed by synteny analysis with MultiContigView tool in the Ensembl genome browser. RESULTS: By integrated analysis of three recently published DNA microarray studies with human tissue, thirty-eight genes were found to be differentially expressed in the same direction in fibroid compared to adjacent uterine myometrium by at least two research groups. Among these genes, twelve with rat orthologs were identified as estrogen-regulated from our array study investigating uterine expression in ovariectomized rats treated with estrogen. Functional and pathway analyses of the twelve genes suggested multiple molecular mechanisms for estrogen-dependent cell survival and tumor growth. Firstly, estrogen increased expression of the anti-apoptotic PCP4 gene and suppressed the expression of growth inhibitory receptors PTGER3 and TGFBR2. Secondly, estrogen may antagonize PPARγ signaling, thought to inhibit fibroid growth and survival, at two points in the PPAR pathway: 1) through increased ANXA1 gene expression which can inhibit phospholipase A2 activity and in turn decrease arachidonic acid synthesis, and 2) by decreasing L-PGDS expression which would reduce synthesis of PGJ2, an endogenous ligand for PPARγ. Lastly, estrogen affects retinoic acid (RA) synthesis and mobilization by regulating expression of CRABP2 and ALDH1A1. RA has been shown to play a significant role in the development of uterine fibroids in an animal model. CONCLUSION: Integrated analysis of multiple array datasets revealed twelve human and rat ortholog genes that were differentially expressed in human uterine fibroids and transcriptionally responsive to estrogen in the rat uterus. Functional and pathway analysis of these genes suggest multiple potential molecular mechanisms for the poorly understood estrogen-dependent growth of uterine fibroids. Fully understanding the exact molecular interactions among these gene products requires further study to validate their roles in uterine fibroids. This work provides new avenues of study which could influence the future direction of therapeutic intervention for the disease

Inhibition of H3K9 Methyltransferase G9a/GLP Prevents Ototoxic and Ongoing Hair Cell Death

Sensorineural hearing loss (SNHL) is one of the most common sensory defects in humans. Considering the susceptibility of hair cells to many vulnerabilities and the important role of histone modifications in cell development and injury, we examined the roles of G9a/GLP in hair cell protection by using pharmacological inhibitors. We observed a transient increase of H3K9me2 upon hair cell damage in organotypic cochlear cultures. Treatment with BIX01294 or UNC0638, specific inhibitors to G9a/GLP, prevented hair cell loss accompanying the reduced level of H3K9me2. Using fluorescent labelling, we demonstrated that BIX01294 does not interfere with aminoglycoside uptake, indicating the otoprotective role of BIX01294 does not likely function through the regulation of aminoglycoside absorption. Moreover, local delivery of BIX01294 prevented neomycin-induced in vivo auditory hair cell loss in the organ of Corti in a mouse mode, as measured by histological and functional auditory brainstem response (ABR) analyses. Transcritptome profiling analysis indicated that G9a/GLP inhibition may up-regulate the expression of to Pcdh8 and Insm1 execute its otoprotection roles. These data revealed a novel role of histone modification in otoprotection. The reduced susceptibility of the hair cells to stress by the downregulation of H3K9me2 may be of potential therapeutic value for those suffering from SNHL

TET-catalyzed 5-methylcytosine hydroxylation is dynamically regulated by metabolites

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