参考文献

A murine leukocyte surface glycoprotein (Mr = 95 000) has been defined by means of xenogeneic monoclonal antibodies. In normal hematopoietic tissues,the glycoprotein is found in highest amounts in the bone marrow. Flow cytometric analysis shows that essentially all bone-marrow cells express the glycoprotein and that it is a major component of a subpopulation of cells containing predominantly granulocytic precursors. In contrast,only about 5 percent of thymocytes express sufficient glycoprotein to be detected by flow cytometric analysis,although under stringent conditions up to 20 percent of thymocytes are susceptible to complement-mediated cytotoxicity using a monoclonal antibody against the glycoprotein. Functional assays showed that both prothymocytes and colony forming unit-spleen express the glycoprotein which is broadly distributed on murine hematopoietic tumor cell lines. However,although some Thy-1+ (T) cell lymphomas express large amounts of the glycoprotein,others do not express detectable quantities of the molecule. The glycoprotein is not restricted to hematopoietic cells and can be detected on lung,kidney,brain,and liver as well as cultured fibroblasts. Monoclonal antibodies against the glycoprotein cross-react with an antigen present on human cells. As described in the accompanying paper,the glycoprotein exists in two antithetical allelic forms and we show that it is identical to a polymorphic surface molecule independently characterized by Colombatti and co-workers. View Publication

Epithelial tissues sheath organs and electro-mechanically regulate ion and water transport to regulate development,homeostasis,and hydrostatic organ pressure. Here,we demonstrate how external electrical stimulation allows us to control these processes in living tissues. Specifically,we electrically stimulate hollow,3D kidneyoids and gut organoids and find that physiological-strength electrical stimulation of ? 5 - 10 V/cm powerfully inflates hollow tissues; a process we call electro-inflation. Electro-inflation is mediated by increased ion flux through ion channels/transporters and triggers subsequent osmotic water flow into the lumen,generating hydrostatic pressure that competes against cytoskeletal tension. Our computational studies suggest that electro-inflation is strongly driven by field-induced ion crowding on the outer surface of the tissue. Electrically stimulated tissues also break symmetry in 3D resulting from electrotaxis and affecting tissue shape. The ability of electrical cues to regulate tissue size and shape emphasizes the role and importance of the electrical micro-environment for living tissues. Electrical stimulation of hollow,3D kidney tissues causes these tissues to inflate and change shape. The authors call this process electro-inflation and connect it to electricity driving ions into the center of the tissues,causing water to follow by osmosis. View Publication

Mitochondrial dysfunction has been demonstrated to result in premature aging due to its effects on stem cells. Nevertheless,a full understanding of the role of mitochondrial bioenergetics through differentiation is still lacking. Here we show the bioenergetics profile of human stem cells of embryonic origin differentiating along the hepatic lineage. Our study reveals especially the transition between hepatic specification and hepatic maturation as dependent on mitochondrial respiration and demonstrates that even though differentiating cells are primarily dependent on glycolysis until induction of hepatocyte maturation,oxidative phosphorylation is essential at all stages of differentiation. View Publication

Primary systemic amyloidosis (AL) is a rare monoclonal plasma cell (PC) disorder characterized by the deposition of misfolded immunoglobulin (Ig) light chains (LC) in vital organs throughout the body. To our knowledge,no cell lines have ever been established from AL patients. Here we describe the establishment of the ALMC-1 and ALMC-2 cell lines from an AL patient. Both cell lines exhibit a PC phenotype and display cytokine-dependent growth. Using a comprehensive genetic approach,we established the genetic relationship between the cell lines and the primary patient cells,and we were also able to identify new genetic changes accompanying tumor progression that may explain the natural history of this patient's disease. Importantly,we demonstrate that free lambda LC secreted by both cell lines contained a beta structure and formed amyloid fibrils. Despite absolute Ig LC variable gene sequence identity,the proteins show differences in amyloid formation kinetics that are abolished by the presence of Na(2)SO(4). The formation of amyloid fibrils from these naturally secreting human LC cell lines is unprecedented. Moreover,these cell lines will provide an invaluable tool to better understand AL,from the combined perspectives of amyloidogenic protein structure and amyloid formation,genetics,and cell biology. View Publication

AIMS Pifithrin α (PFTα),an inhibitor of the p53 protein,is regarded as a lead compound for cancer and neurodegenerative disease therapy. There is some evidence that this compound activates the aryl hydrocarbon receptor (AhR) in a complete independent way of the p53 inhibition and that it is easily converted to its condensation product pifithrin β (PFTβ). The aim of this study was to explore the ability of PFTα and of PFTβ to induce a variety of AhR mediated processes. MAIN METHODS Computational analysis using quantum chemical calculations and chemical analysis have been used to study the conformation of the compounds as well as the cyclization reaction. The AhR mediated processes of these compounds have been studied in a rainbow trout cell line (RTG-2) and in a rat hepatoma cell line (H4IIE). KEY FINDINGS PFTα molecule could not take a planar conformation required for AhR activation whereas PFTβ showed a conformation similar to those of the prototypical AhR ligand β-naphthoflavone. In both cell lines,PFTα and PFTβ provoked different responses related with AhR activation. However,when cyclization of PFTα to PFTβ was hampered by acetylation of the exocyclic nitrogen,all these responses were not observed. These results lead to the conclusion that the activation of the AhR is probably caused by PFTβ instead of PFTα. SIGNIFICANCE Since PFTα is a promising compound for the development of new pharmaceuticals inhibiting p53,the chemical instability of this compound as well as the capacity of its transformation product should be taken into account. View Publication

PURPOSE: The Ras-Raf-mitogen-activated protein kinase kinase (MEK) pathway is overactive in many human cancers and is thus a target for novel therapeutics. We have developed a highly potent and selective inhibitor of MEK1/2. The purpose of these studies has been to show the biological efficacy of ARRY-142886 (AZD6244) in enzymatic,cellular,and animal models. EXPERIMENTAL DESIGN: The ability of ARRY-142886 to inhibit purified MEK1 as well as other kinases was evaluated. Its effects on extracellular signal-regulated kinase (ERK) phosphorylation and proliferation in several cell lines were also determined. Finally,the inhibitor was tested in HT-29 (colorectal) and BxPC3 (pancreatic) xenograft tumor models. RESULTS: The IC(50) of ARRY-142886 was determined to be 14 nmol/L against purified MEK1. This activity is not competitive with ATP,which is consistent with the high specificity of compound for MEK1/2. Basal and epidermal growth factor-induced ERK1/2 phosphorylation was inhibited in several cell lines as well as 12-O-tetradecanoylphorbol-13-acetate-induced ERK1/2 phosphorylation in isolated peripheral blood mononuclear cells. Treatment with ARRY-142886 resulted in the growth inhibition of several cell lines containing B-Raf and Ras mutations but had no effect on a normal fibroblast cell line. When dosed orally,ARRY-142886 was capable of inhibiting both ERK1/2 phosphorylation and growth of HT-29 xenograft tumors in nude mice. Tumor regressions were also seen in a BxPC3 xenograft model. In addition,tumors remained responsive to growth inhibition after a 7-day dosing holiday. CONCLUSIONS: ARRY-142886 is a potent and selective MEK1/2 inhibitor that is highly active in both in vitro and in vivo tumor models. This compound is currently being investigated in clinical studies. View Publication

The phosphatidylinositide 3-kinase pathway is frequently deregulated in human cancers and inhibitors offer considerable therapeutic potential. We previously described the promising tricyclic pyridofuropyrimidine lead and chemical tool compound PI-103. We now report the properties of the pharmaceutically optimized bicyclic thienopyrimidine derivatives PI-540 and PI-620 and the resulting clinical development candidate GDC-0941. All four compounds inhibited phosphatidylinositide 3-kinase p110alpha with IC(50) textless or = 10 nmol/L. Despite some differences in isoform selectivity,these agents exhibited similar in vitro antiproliferative properties to PI-103 in a panel of human cancer cell lines,with submicromolar potency in PTEN-negative U87MG human glioblastoma cells and comparable phosphatidylinositide 3-kinase pathway modulation. PI-540 and PI-620 exhibited improvements in solubility and metabolism with high tissue distribution in mice. Both compounds gave improved antitumor efficacy over PI-103,following i.p. dosing in U87MG glioblastoma tumor xenografts in athymic mice,with treated/control values of 34% (66% inhibition) and 27% (73% inhibition) for PI-540 (50 mg/kg b.i.d.) and PI-620 (25 mg/kg b.i.d.),respectively. GDC-0941 showed comparable in vitro antitumor activity to PI-103,PI-540,and PI-620 and exhibited 78% oral bioavailability in mice,with tumor exposure above 50% antiproliferative concentrations for textgreater8 hours following 150 mg/kg p.o. and sustained phosphatidylinositide 3-kinase pathway inhibition. These properties led to excellent dose-dependent oral antitumor activity,with daily p.o. dosing at 150 mg/kg achieving 98% and 80% growth inhibition of U87MG glioblastoma and IGROV-1 ovarian cancer xenografts,respectively. Together,these data support the development of GDC-0941 as a potent,orally bioavailable inhibitor of phosphatidylinositide 3-kinase. GDC-0941 has recently entered phase I clinical trials. View Publication

IL-3,IL-5,and GM-CSF are related hematopoietic cytoines that are important for allergic inflammation. The receptors for human IL-5,IL-3,and GM-CSF are members of the hematopoietin receptor superfamily and are comprised of a cytokine-specific alpha chain and the common beta chain that is shared among these cytokines for signaling. Each of these cytokines contributes to the differentiation and function of leukocyte subpopulations and have clinical importance in protective immunity and in the pathophysiology of a spectrum of immunologic diseases that are as diverse as allergy and asthma,pulmonary alveolar proteinosis,neurodegenerative diseases,and malignancies. Delineating the biology of these cytokines is enabling the development of new strategies for diagnosing and treating these diseases and modulating immune responses. View Publication

Myocardial infarction is a leading cause of mortality and morbidity worldwide,and current treatments fail to address the underlying scarring and cell loss,which is a major cause of heart failure after infarction. The novel strategy,therapeutic angiogenesis and/or vasculogenesis with endothelial progenitor cells transplantation holds great promise to increase blood flow in ischemic areas,thus rebuild the injured heart and reverse the heart failure. Given the potential of self-renewal and differentiation into virtually all cell types,human embryonic stem cells (hESCs) may provide an alternate source of therapeutic cells by allowing the derivation of large numbers of endothelial cells for therapeutic angiogenesis and/or vasculogenesis of ischemic heart diseases. Moreover,to fully understand the fate of implanted hESCs or hESC derivatives,investigators need to monitor the motility of cells in living animals over time. In this chapter,we describe the application of bioluminescence reporter gene imaging to track the transplanted hESC-derived endothelial cells for treatment of myocardial infarction. The technology of inducing endothelial cells from hESCs will also be discussed. View Publication

Human embryonic stem (hES) cells have a potential use for the repair and regeneration of injured tissues. However,teratoma formation can be a major obstacle for hES-mediated cell therapy. Therefore,tracking the fate and function of transplanted hES cells with noninvasive imaging could be valuable for a better understanding of the biology and physiology of teratoma formation. In this study,hES cells were stably transduced with a double fusion reporter gene consisting of firefly luciferase and enhanced green fluorescent protein. Following bioluminescence imaging and histology,we demonstrated that engraftment of hES cells was followed by dramatically increasing signaling and led to teratoma formation confirmed by histology. Studies of the angiogenic processes within teratomas revealed that their vasculatures were derived from both differentiated hES cells and host. Moreover,FACS analysis showed that teratoma cells derived from hES cells expressed high levels of CD56 and SSEA-4,and the subcultured SSEA-4(+) cells showed a similar cell surface marker expression pattern when compared to undifferentiated hES cells. We report here for the first time that SSEA-4(+) cells derived from teratoma exhibited multipotency,retained their differentiation ability in vivo as confirmed by their differentiation into representative three germ layers. View Publication