参考文献

Cell surface biomarkers have been applied to discriminate pluripotent human embryonic stem cells and induced pluripotent stem cells from differentiated cells. Here,we demonstrate that a recombinant lectin probe,rBC2LCN,a new tool for fluorescence-based imaging and flow cytometry analysis of pluripotent stem cells,is an alternative to conventional pluripotent maker antibodies. Live or fixed colonies of both human embryonic stem cells and induced pluripotent stem cells were visualized in culture medium containing fluorescent dye-labeled rBC2LCN. Fluorescent dye-labeled rBC2LCN was also successfully used to separate live pluripotent stem cells from a mixed cell population by flow cytometry. textcopyright 2013 Elsevier Inc. View Publication

The use of human pluripotent stem cells (hPSCs) in cell therapy is hindered by the tumorigenic risk from residual undifferentiated cells. Here we performed a high-throughput screen of over 52,000 small molecules and identified 15 pluripotent cell-specific inhibitors (PluriSIns),nine of which share a common structural moiety. The PluriSIns selectively eliminated hPSCs while sparing a large array of progenitor and differentiated cells. Cellular and molecular analyses demonstrated that the most selective compound,PluriSIn 1,induces ER stress,protein synthesis attenuation,and apoptosis in hPSCs. Close examination identified this molecule as an inhibitor of stearoyl-coA desaturase (SCD1),the key enzyme in oleic acid biosynthesis,revealing a unique role for lipid metabolism in hPSCs. PluriSIn 1 was also cytotoxic to mouse blastocysts,indicating that the dependence on oleate is inherent to the pluripotent state. Finally,application of PluriSIn 1 prevented teratoma formation from tumorigenic undifferentiated cells. These findings should increase the safety of hPSC-based treatments. ?? 2013 Elsevier Inc. View Publication

The importance of anticancer stem cell research for breast cancer lies in the possibility of providing new approaches for an improved understanding of anticancer activity and cancer treatment. In this study,we demonstrated that the preclinical therapeutic efficacy of combining the multikinase inhibitor sorafenib with radiation was more effective in hypoxia-exposed breast cancer stem cells. We assessed cell viability and Annexin V to evaluate the combined effect of sorafenib and radiation following exposure to hypoxia. Our results showed that the synergistic cytotoxicity increased tumor cell apoptosis significantly and reduced cell proliferation in MDA-MB-231 and MCF-7 cells under hypoxic conditions compared to sorafenib or radiation alone in vitro. Additionally,the combined treatment induced G2/M cell cycle arrest. Notably,the combination of sorafenib and radiation eliminated CD44+CD24-/low cells preferentially,which highly expressed hypoxia-inducible factor (HIF)-1$$ and effectively inhibited primary and secondary mammosphere formation in MDA-MB-231 cells. A combined effect on MDA-MB‑231 cells in response to hypoxia was shown by inhibiting angiogenesis and metastasis by suppression of HIF-1$$ and matrix metalloproteinase-2 (MMP-2). Collectively,these results indicate that the efficacy of sorafenib combined with radiation for treating human breast cancer cells is synergistic and suggest a new therapeutic approach to prevent breast cancer progression by eliminating breast cancer stem cells. View Publication

Hearing loss due to damage to auditory hair cells is normally irreversible because mammalian hair cells do not regenerate. Here,we show that new hair cells can be induced and can cause partial recovery of hearing in ears damaged by noise trauma,when Notch signaling is inhibited by a γ-secretase inhibitor selected for potency in stimulating hair cell differentiation from inner ear stem cells in vitro. Hair cell generation resulted from an increase in the level of bHLH transcription factor Atoh1 in response to inhibition of Notch signaling. In vivo prospective labeling of Sox2-expressing cells with a Cre-lox system unambiguously demonstrated that hair cell generation resulted from transdifferentiation of supporting cells. Manipulating cell fate of cochlear sensory cells in vivo by pharmacological inhibition of Notch signaling is thus a potential therapeutic approach to the treatment of deafness. View Publication

Wnt/$$-catenin signalling has been suggested to be active in basal-like breast cancer. However,in highly aggressive metastatic triple-negative breast cancers (TNBC) the role of $$-catenin and the underlying mechanism(s) for the aggressiveness of TNBC remain unknown. We illustrate that WNT10B induces transcriptionally active $$-catenin in human TNBC and predicts survival-outcome of patients with both TNBC and basal-like tumours. We provide evidence that transgenic murine Wnt10b-driven tumours are devoid of ER$$,PR and HER2 expression and can model human TNBC. Importantly,HMGA2 is specifically expressed during early stages of embryonic mammogenesis and absent when WNT10B expression is lost,suggesting a developmentally conserved mode of action. Mechanistically,ChIP analysis uncovered that WNT10B activates canonical $$-catenin signalling leading to up-regulation of HMGA2. Treatment of mouse and human triple-negative tumour cells with two Wnt/$$-catenin pathway modulators or siRNA to HMGA2 decreases HMGA2 levels and proliferation. We demonstrate that WNT10B has epistatic activity on HMGA2,which is necessary and sufficient for proliferation of TNBC cells. Furthermore,HMGA2 expression predicts relapse-free-survival and metastasis in TNBC patients. View Publication

Mammalian target of rapamycin (mTOR) is essential in controlling several cellular functions. This pathway is dysregulated in keloid disease (KD). KD is a common fibroproliferative dermal lesion with an ill-defined treatment strategy. KD demonstrates excessive matrix deposition,angiogenesis,and inflammatory cell infiltration. In KD,both total and phosphorylated forms of mTOR and p70(S6K)(Thr421/Ser424) are upregulated. Therefore,the aim of this study was to investigate adenosine triphosphate-competitive inhibitors of mTOR kinase previously unreported in keloid and their comparative efficacy with Rapamycin. Here,we present two mTOR kinase inhibitors,KU-0063794 and KU-0068650,that target both mTORC1 and mTORC2 signaling. Treatment with either KU-0063794 or KU-0068650 resulted in complete suppression of Akt,mTORC1,and mTORC2,and inhibition of keloid cell spreading,proliferation,migration,and invasive properties at a very low concentration (2.5 μmol l(-1)). Both KU-0063794 and KU-0068650 significantly (Ptextless0.05) inhibited cell cycle regulation and HIF1-α expression compared with that achieved with Rapamycin alone. In addition,both compounds induced shrinkage and growth arrest in KD,associated with the inhibition of angiogenesis,induction of apoptosis,and reduction in keloid phenotype-associated markers. In contrast,Rapamycin induced minimal antitumor activity. In conclusion,potent dual mTORC1 and mTORC2 inhibitors display therapeutic potential for the treatment of KD. View Publication

Cultures of human embryonic stem cell typically rely on protein matrices or feeder cells to support attachment and growth,while mechanical,enzymatic or chemical cell dissociation methods are used for cellular passaging. However,these methods are ill defined,thus introducing variability into the system,and may damage cells. They also exert selective pressures favouring cell aneuploidy and loss of differentiation potential. Here we report the identification of a family of chemically defined thermoresponsive synthetic hydrogels based on 2-(diethylamino)ethyl acrylate,which support long-term human embryonic stem cell growth and pluripotency over a period of 2-6 months. The hydrogels permitted gentle,reagent-free cell passaging by virtue of transient modulation of the ambient temperature from 37 to 15 °C for 30 min. These chemically defined alternatives to currently used,undefined biological substrates represent a flexible and scalable approach for improving the definition,efficacy and safety of human embryonic stem cell culture systems for research,industrial and clinical applications. View Publication

It has been 50 years since cellular senescence was first described in human diploid fibroblasts (HDFs),yet its mechanism as well as its physiological and clinical implications are still not fully appreciated. Recent progress suggests that cellular senescence is a collective phenotype,composed of complex networks of effector programs. The balance and quality within the effector network varies depending on the cell type,the nature of the stress as well as the context. Therefore,understanding each of these effectors in the context of the whole network will be necessary in order to fully understand senescence as a whole. Furthermore,searching for new effector programs of senescence will help to define this heterogeneous and complex phenotype according to cellular contexts. View Publication

Familial hypertrophic cardiomyopathy (HCM) is a prevalent hereditary cardiac disorder linked to arrhythmia and sudden cardiac death. While the causes of HCM have been identified as genetic mutations in the cardiac sarcomere,the pathways by which sarcomeric mutations engender myocyte hypertrophy and electrophysiological abnormalities are not understood. To elucidate the mechanisms underlying HCM development,we generated patient-specific induced pluripotent stem cell cardiomyocytes (iPSC-CMs) from a ten-member family cohort carrying a hereditary HCM missense mutation (Arg663His) in the MYH7 gene. Diseased iPSC-CMs recapitulated numerous aspects of the HCM phenotype including cellular enlargement and contractile arrhythmia at the single-cell level. Calcium (Ca2+) imaging indicated dysregulation of Ca2+ cycling and elevation in intracellular Ca2+ ([Ca2+] i) are central mechanisms for disease pathogenesis. Pharmacological restoration of Ca2+ homeostasis prevented development of hypertrophy and electrophysiological irregularities. We anticipate that these findings will help elucidate the mechanisms underlying HCM development and identify novel therapies for the disease. textcopyright 2013 Elsevier Inc. View Publication

Breast cancer is a heterogenous disease,composed of tumour cells with differing gene expressions and phenotypes. Very few antigens have been identified and a better understanding of tumour initiating-cells as targets for therapy is critically needed. Recently,a rare subpopulation of cells within tumours has been described with the ability to: (i) initiate and sustain tumour growth; (ii) resist traditional therapies and allow for secondary tumour dissemination; and (iii) display some of the characteristics of stem cells such as self-renewal. These cells are termed tumour-initiating cells or cancer stem cells,or alternatively,in the case of breast cancer,breast cancer stem cells. Previous studies have demonstrated that breast cancer stem cells can be enriched for in tumoursphere" culture. Proteomics represents a novel way to investigate protein expression between cells. We hypothesise that characterisation of the proteome of the breast cancer line MCF-7 tumourspheres compared to adherent/differentiated cells identifies proteins of novel interest for further isolating or targeting breast cancer stem cells. We present evidence that: (i) the proteome of adherent cells is different to the proteome of cells grown in sphere medium from either early passage (passage 2) or late passage (passage 5) spheres; (ii) that spheres are enriched in expression of a variety of tumour-relevant proteins (including MUC1 and Galectin-3); and (iii) that targeting of one of these identified proteins (galectin-3) using an inhibitor (N-acetyllactosamine) decreases sphere formation/self-renewal of MCF-7 cancer stem cells in vitro and tumourigenicity in vivo. Hence� View Publication