[Show abstract][Hide abstract] ABSTRACT: Abstract Alterations of DNA methylation occur during the course of both stem cell development and tumorigenesis. We present a novel strategy that can be used to stratify glioblastoma multiforme (GBM) patients through the epigenetic states of genes associated with human embryonic stem cell (hESC) identity in order to 1) assess linkages between the methylation signatures of these stem cell genes and survival of GBM patients, and 2) delineate putative mechanisms leading to poor prognosis in some patient subgroups. A DNA methylation signature was established for stratifying GBM patients into several hESC methylator subgroups. The hESC methylator-negative phenotype has demonstrated poor survival and upregulation of glioma stem cell (GSC) markers, and is enriched in one of the previously defined transcriptomic phenotypes-the mesenchymal phenotype. We further identified a refined signature of 36 genes as the gene panel, including SOX2, POU3F2, FGFR2, GAP43, NTRK2, NTRK3, and NKX2-2, which are highly enriched in the nervous system. Both signatures outperformed the O6-methylguanine-DNA methyltransferase (MGMT) methylation test in predicting patient's outcome. These findings were also validated through an independent dataset of patients. Furthermore, through statistical analyses, both signatures were examined significantly. Hypomethylation of hESC-associated genes predicted poorer clinical outcome in GBM, supporting the idea that epigenetic activation of stem cell genes contributes to GBM aggression. The gene panel presented herein may be developed into clinical assays for patient stratification and future personalized medicine interventions.
Omics: a journal of integrative biology 03/2014; 18(5). DOI:10.1089/omi.2013.0084 · 2.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In immune responses, activated T cells migrate to B-cell follicles and develop into follicular T-helper (TFH) cells, a recently identified subset of CD4(+) T cells specialized in providing help to B lymphocytes in the induction of germinal centres. Although Bcl6 has been shown to be essential in TFH-cell function, it may not regulate the initial migration of T cells or the induction of the TFH program, as exemplified by C-X-C chemokine receptor type 5 (CXCR5) upregulation. Here we show that expression of achaete-scute homologue 2 (Ascl2)-a basic helix-loop-helix (bHLH) transcription factor-is selectively upregulated in TFH cells. Ectopic expression of Ascl2 upregulates CXCR5 but not Bcl6, and downregulates C-C chemokine receptor 7 (CCR7) expression in T cells in vitro, as well as accelerating T-cell migration to the follicles and TFH-cell development in vivo in mice. Genome-wide analysis indicates that Ascl2 directly regulates TFH-related genes whereas it inhibits expression of T-helper cell 1 (TH1) and TH17 signature genes. Acute deletion of Ascl2, as well as blockade of its function with the Id3 protein in CD4(+) T cells, results in impaired TFH-cell development and germinal centre response. Conversely, mutation of Id3, known to cause antibody-mediated autoimmunity, greatly enhances TFH-cell generation. Thus, Ascl2 directly initiates TFH-cell development.
[Show abstract][Hide abstract] ABSTRACT: Reversible ubiquitin modification of cell signaling molecules has emerged as a critical mechanism by which cells respond to extracellular stimuli. Although ubiquitination of TGF-β-activated kinase 1 (TAK1) is critical for NF-κB activation in T cells, the regulation of its deubiquitination is unclear. We show that USP18, which was previously reported to be important in regulating type I interferon signaling in innate immunity, regulates T cell activation and T helper 17 (Th17) cell differentiation by deubiquitinating the TAK1-TAB1 complex. USP18-deficient T cells are defective in Th17 differentiation and Usp18(-/-) mice are resistant to experimental autoimmune encephalomyelitis (EAE). In response to T cell receptor engagement, USP18-deficient T cells exhibit hyperactivation of NF-κB and NFAT and produce increased levels of IL-2 compared with the wild-type controls. Importantly, USP18 is associated with and deubiquitinates the TAK1-TAB1 complex, thereby restricting expression of IL-2. Our findings thus demonstrate a previously uncharacterized negative regulation of TAK1 activity during Th17 differentiation, suggesting that USP18 may be targeted to treat autoimmune diseases.
Journal of Experimental Medicine 07/2013; 210(8). DOI:10.1084/jem.20122327 · 12.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: E14.Tg2a mouse embryonic stem (mES) cells are a widely used host in gene trap and gene targeting techniques. Molecular characterization of host cells will provide background information for a better understanding of functions of the knockout genes. Using a highly selective glycopeptide-capture approach but ordinary liquid chromatography coupled mass spectrometry (LC-MS), we characterized the N-glycoproteins of E14.Tg2a cells and analyzed the close relationship between the obtained N-glycoproteome and cell-surface proteomes. Our results provide a global view of cell surface protein molecular properties, in which receptors seem to be much more diverse but lower in abundance than transporters on average. In addition, our results provide a systematic view of the E14.Tg2a N-glycosylation, from which we discovered some striking patterns, including an evolutionarily preserved and maybe functionally selected complementarity between N-glycosylation and the transmembrane structure in protein sequences. We also observed an environmentally influenced N-glycosylation pattern among glycoenzymes and extracellular matrix proteins. We hope that the acquired information enhances our molecular understanding of mES E14.Tg2a as well as the biological roles played by N-glycosylation in cell biology in general.
PLoS ONE 02/2013; 8(2):e55722. DOI:10.1371/journal.pone.0055722 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: T follicular helper cells (Tfh cells) play a pivotal role in germinal center reactions, which require B cell lymphoma 6 (Bcl6) transcription factor. To analyze their relationships with other effector T cell lineages and their stability in vivo, we developed and analyzed a new Bcl6 reporter mouse alone or together with other lineage reporter systems. Assisted with genome-wide transcriptome analysis, we show substantial plasticity of T cell differentiation in the early phase of immune response. At this stage, CXCR5 appears to be expressed in a Bcl6-independent manner. Once Bcl6 is highly expressed, Tfh cells can persist in vivo and some of them develop into memory cells. Together, our results indicate Bcl6 as a bona fide marker for Tfh polarized program.
Journal of Experimental Medicine 09/2012; 209(10):1841-52. DOI:10.1084/jem.20120219 · 12.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The development and application of systems strategies to biology and disease are transforming medical research and clinical practice in an unprecedented rate. In the foreseeable future, clinicians, medical researchers, and ultimately the consumers and patients will be increasingly equipped with a deluge of personal health information, e.g., whole genome sequences, molecular profiling of diseased tissues, and periodic multi-analyte blood testing of biomarker panels for disease and wellness. The convergence of these practices will enable accurate prediction of disease susceptibility and early diagnosis for actionable preventive schema and personalized treatment regimes tailored to each individual. It will also entail proactive participation from all major stakeholders in the health care system. We are at the dawn of predictive, preventive, personalized, and participatory (P4) medicine, the fully implementation of which requires marrying basic and clinical researches through advanced systems thinking and the employment of high-throughput technologies in genomics, proteomics, nanofluidics, single-cell analysis, and computation strategies in a highly-orchestrated discipline we termed translational systems medicine.
[Show abstract][Hide abstract] ABSTRACT: A grand challenge impeding optimal treatment outcomes for patients with cancer arises from the complex nature of the disease: the cellular heterogeneity, the myriad of dysfunctional molecular and genetic networks as results of genetic (somatic) and environmental perturbations. Systems biology, with its holistic approach to understanding fundamental principles in biology, and the empowering technologies in genomics, proteomics, single-cell analysis, microfluidics and computational strategies, enables a comprehensive approach to medicine, which strives to unveil the pathogenic mechanisms of diseases, identify disease biomarkers and begin thinking about new strategies for drug target discovery. The integration of multidimensional high-throughput 'omics' measurements from tumour tissues and corresponding blood specimens, together with new systems strategies for diagnostics, enables the identification of cancer biomarkers that will enable presymptomatic diagnosis, stratification of disease, assessment of disease progression, evaluation of patient response to therapy and the identification of reoccurrences. Whilst some aspects of systems medicine are being adopted in clinical oncology practice through companion molecular diagnostics for personalized therapy, the mounting influx of global quantitative data from both wellness and diseases is shaping up a transformational paradigm in medicine we termed 'predictive', 'preventive', 'personalized', and 'participatory' (P4) medicine, which requires new strategies, both scientific and organizational, to enable bringing this revolution in medicine to patients and to the healthcare system. P4 medicine will have a profound impact on society - transforming the healthcare system, turning around the ever escalating costs of healthcare, digitizing the practice of medicine and creating enormous economic opportunities for those organizations and nations that embrace this revolution.
Journal of Internal Medicine 12/2011; 271(2):111-21. DOI:10.1111/j.1365-2796.2011.02498.x · 6.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: CD133, a member of the prominin family, is found in a variety of tissues with at least three variants. The function of CD133 is not well understood, but its expression is subject to changes in the microenvironment cues including bioenergetic stress. Knockout of CD133 does not affect renewal, but mammary gland branching. A point mutation of CD133 (R733C) leads to retinal disorder. CD133 is found in embryonic stem cells, normal tissue stem cells, stem cell niches, and circulating endothelial progenitors as well as cancer stem cells. Maintenance of stemness in cancer may be attributable to asymmetric cell division in association with a set of embryonic expression signatures in CD133+ tumor cells. CD133 could enrich cancer stem cells, which are associated with chemo- and radiation resistance phenotype. High CD133 is associated with poor survival in a variety of solid tumors, including lung, colon, prostate, etc. Monitoring CD133+ cells in peripheral blood, and targeting CD133 in cancer, may further predict and improve the clinical outcomes.
Current Colorectal Cancer Reports 12/2011; 7(4):253-259. DOI:10.1007/s11888-011-0106-1
[Show abstract][Hide abstract] ABSTRACT: Cancer cells are heterogeneous and, it has been proposed, fall into at least two classes: the tumor-initiating cancer stem cells (CSC) and the more differentiated tumor cells. The transmembrane protein CD133 has been widely used to isolate putative CSC populations in several cancer types, but its validity as a CSC marker and hence its clinical ramifications remain controversial. Here, we conducted transcriptomic profiling of sorted CD133(+) and CD133(-) cells from human glioblastoma multiforme (GBM) and, by subtractive analysis, established a CD133 gene expression signature composed of 214 differentially expressed genes. Extensive computational comparisons with a compendium of published gene expression profiles reveal that the CD133 gene signature transcriptionally resembles human ES cells and in vitro cultured GBM stem cells, and this signature successfully distinguishes GBM from lower-grade gliomas. More importantly, the CD133 gene signature identifies an aggressive subtype of GBM seen in younger patients with shorter survival who bear excessive genomic mutations as surveyed through the Cancer Genome Atlas Network GBM mutation spectrum. Furthermore, the CD133 gene signature distinguishes higher-grade breast and bladder cancers from their lower-grade counterparts. Our systematic analysis provides molecular and genetic support for the stem cell-like nature of CD133(+) cells and an objective means for evaluating cancer aggressiveness.
Proceedings of the National Academy of Sciences 01/2011; 108(4):1591-6. DOI:10.1073/pnas.1018696108 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Toll-like receptors (TLRs) have previously been shown to play critical roles in the activation of innate immunity. Here, we describe that T cell expression of TLR2 regulates T helper 17 (Th17) cell responses. Stimulation with TLR2 agonists promoted Th17 differentiation in vitro and led to more robust proliferation and Th17 cytokine production. Using the experimental autoimmune encephalomyelitis (EAE) model, we found that TLR2 regulated Th17 cell-mediated autoimmunity in vivo and that loss of TLR2 in CD4(+) T cells dramatically ameliorated EAE. This study thus reveals a critical role of a TLR in the direct regulation of adaptive immune response and pathogenesis of autoimmune diseases.
[Show abstract][Hide abstract] ABSTRACT: Epigenetic inactivation of tumor suppressor genes is common in human cancer. Using a large-scale whole-genome approach in an earlier study, the authors identified epigenetically silenced genes with potential tumor suppressor function in glioblastoma (GBM). Three genes identified in this analysis-DKK1, SFRP1, and WIF1-are potent inhibitors of the Wnt signal transduction pathway. Here, the authors confirm decreased expression of these genes in GBM tumor tissue samples relative to nontumor brain tissue samples using real-time PCR. They then show that expression of all 3 genes is restored in T98 GBM cells by treatment with the histone deacetylase inhibitor Trichostatin A (TSA), but only DKK1 expression is restored by treatment with the demethylating agent 5-azacytidine. Bisulfite sequencing did not reveal significant methylation in the promoter region of DKK1, whereas histone acetylation and chromatin accessibility increased significantly for all 3 genes after TSA treatment. Ectopic expression of DKK1 significantly reduces colony formation and increases chemotherapy-induced apoptosis in T98 cells. Ectopic expression of the canonical Wnt pathway inhibitors WIF1 and SFRP1 shows a relative lack of response. Chronic Wnt3a stimulation only partially reverses growth suppression after DKK1 reexpression, whereas a specific inhibitor of the JNK pathway significantly reverses the effect of DKK1 reexpression on colony formation and apoptosis in T98 cells. These results support a potential growth-suppressive function for epigenetically silenced DKK1 in GBM and suggest that DKK1 restoration could modulate Wnt signaling through both canonical and noncanonical pathways.
Genes & cancer 01/2010; 1(1):81-90. DOI:10.1177/1947601909356103
[Show abstract][Hide abstract] ABSTRACT: Cancer stem cells (CSC), also called tumor initiating cells (TIC), are considered to be the origin of replicating malignant tumor cells in a variety of human cancers. Their presence in the tumor may herald malignancy potential, mediate resistance to conventional chemotherapy or radiotherapy, and confer poor survival outcomes. Thus, CSC may serve as critical cellular targets for treatment. The ability to therapeutically target CSC hinges upon identifying their unique cell surface markers and the underlying survival signaling pathways. While accumulating evidence suggests cell-surface antigens (such as CD44, CD133) as CSC markers for several tumor tissues, emerging clinical needs exist for the identification of new markers to completely separate CSC from normal stem cells. Recent studies have demonstrated the critical role of the tumor suppressor PTEN/PI3 kinase pathway in regulating TIC in leukemia, brain, and intestinal tissues. The successful eradication of tumors by therapies targeting CSC will require an in-depth understanding of the molecular mechanisms governing CSC self renewal, differentiation, and escape from conventional therapy. Here we review recent progress from brain tumor and intestinal stem cell research with a focus on the PTEN-Akt-Wnt pathway, and how the components of CSC pathways may serve as biomarkers for diagnosis, prognosis, and therapeutics.
Current Stem Cell Research & Therapy 06/2009; 4(2):147-53. DOI:10.2174/157488809788167373 · 2.21 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Embryonic stem cells (ESCs) are endowed with the ability to generate multiple cell lineages and carry great therapeutic potentials in regenerative medicine. Future application of ESCs in human health and diseases will embark on the delineation of molecular mechanisms that define the biology of ESCs. Here, we discuss how the finite ESC components mediate the intriguing task of brain development and exhibit biomedical potentials to cure diverse neurological disorders. Birth Defects Research (Part C) 87:182–191, 2009.
Birth Defects Research Part C Embryo Today Reviews 06/2009; 87(2):182-91. DOI:10.1002/bdrc.20149 · 2.63 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: T helper (Th) 17 cells have been recently discovered in both mouse and human. Here we show that interleukin-1 (IL-1) signaling on T cells is critically required for the early programming of Th17 cell lineage and Th17 cell-mediated autoimmunity. IL-1 receptor1 expression in T cells, which was induced by IL-6, was necessary for the induction of experimental autoimmune encephalomyelitis and for early Th17 cell differentiation in vivo. Moreover, IL-1 signaling in T cells was required in dendritic cell-mediated Th17 cell differentiation from naive or regulatory precursors and IL-1 synergized with IL-6 and IL-23 to regulate Th17 cell differentiation and maintain cytokine expression in effector Th17 cells. Importantly, IL-1 regulated the expression of the transcription factors IRF4 and RORgammat during Th17 cell differentiation; overexpression of these two factors resulted in IL-1-independent Th17 cell polarization. Our data thus indicate a critical role of IL-1 in Th17 cell differentiation and this pathway may serve as a unique target for Th17 cell-mediated immunopathology.
[Show abstract][Hide abstract] ABSTRACT: T cell activation and tolerance are delicately regulated by costimulatory molecules. Although B and T lymphocyte attenuator (BTLA) has been shown as a negative regulator for T cell activation, its role in peripheral T cell tolerance induction in vivo has not been addressed. In this study, we generated a novel strain of BTLA-deficient mice and used three different models to characterize the function of BTLA in controlling T cell tolerance. In an oral tolerance model, BTLA-deficient mice were found resistant to the induction of T cell tolerance to an oral Ag. Moreover, compared with wild-type OT-II cells, BTLA(-/-) OT-II cells were less susceptible to tolerance induction by a high-dose OVA peptide administered i.v. Finally, BTLA(-/-) OT-I cells caused autoimmune diabetes in RIP-mOVA recipient mice. Our results thus demonstrate an important role for BTLA in the induction of peripheral tolerance of both CD4(+) and CD8(+) T cells in vivo.
The Journal of Immunology 05/2009; 182(8):4516-20. DOI:10.4049/jimmunol.0803161 · 4.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We performed the first genome-wide expression analysis directly comparing the expression profile of highly enriched normal human hematopoietic stem cells (HSC) and leukemic stem cells (LSC) from patients with acute myeloid leukemia (AML). Comparing the expression signature of normal HSC to that of LSC, we identified 3,005 differentially expressed genes. Using 2 independent analyses, we identified multiple pathways that are aberrantly regulated in leukemic stem cells compared with normal HSC. Several pathways, including Wnt signaling, MAP Kinase signaling, and Adherens Junction, are well known for their role in cancer development and stem cell biology. Other pathways have not been previously implicated in the regulation of cancer stem cell functions, including Ribosome and T Cell Receptor Signaling pathway. This study demonstrates that combining global gene expression analysis with detailed annotated pathway resources applied to highly enriched normal and malignant stem cell populations, can yield an understanding of the critical pathways regulating cancer stem cells.
Proceedings of the National Academy of Sciences 03/2009; 106(9):3396-401. DOI:10.1073/pnas.0900089106 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Th17 and regulatory T (Treg) cells play opposite roles in autoimmune diseases. However, the mechanisms underlying their proper migration to inflammatory tissues are unclear. In this study, we report that these two T cell subsets both express CCR6. CCR6 expression in Th17 cells is regulated by TGF-beta and requires two nuclear receptors, RORalpha and RORgamma. Th17 cells also express the CCR6 ligand CCL20, which is induced synergistically by TGF-beta and IL-6, which requires STAT3, RORgamma and IL-21. Th17 cells, by producing CCL20, promote migration of Th17 and Treg cells in vitro in a CCR6-dependent manner. Lack of CCR6 in Th17 cells reduces the severity of experimental autoimmune encephalomyelitis and Th17 and Treg recruitment into inflammatory tissues. Similarly, CCR6 on Treg cells is also important for their recruitment into inflammatory tissues. Our data indicate an important role of CCR6 in Treg and Th17 cell migration.
The Journal of Immunology 01/2009; 181(12):8391-401. DOI:10.4049/jimmunol.181.12.8391 · 4.92 Impact Factor